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Sample records for adjuvant-free murine experimental

  1. Intranasal vaccination with adjuvant-free S. aureus antigens effectively protects mice against experimental sepsis.

    Stegmiller, Nataly Pescinalli; Barcelos, Estevão Carlos; Leal, Janine Miranda; Covre, Luciana Polaco; Donatele, Dirlei Molinari; de Matos Guedes, Herbet Leonel; Cunegundes, Marco Cesar; Rodrigues, Rodrigo Ribeiro; Gomes, Daniel Cláudio Oliviera

    2016-06-24

    Staphylococcus aureus (S. aureus) is a Gram-positive coccal bacterium comprising part of the human skin, nares and gastrointestinal tract normal microbiota. It is also an important cause of nosocomial/community-acquired infections in humans and animals, which can cause a diverse array of infections, including sepsis, which is a progressive systemic inflammation response syndrome that is frequently fatal. The emergence of drug-resistant strains and the high toxicity of the treatments used for these infections point out the need to develop an effective, inexpensive and safe vaccine that can be used prophylactically. In this work, we used an experimental sepsis model to evaluate the effectiveness of whole antigens from S. aureus (SaAg) given by the intranasal route to induce protective immunity against S. aureus infection in mice. BALB/c mice were vaccinated via intranasal or intramuscular route with two doses of SaAg, followed by biocompatibility and immunogenicity evaluations. Vaccinated animals did not show any adverse effects associated with the vaccine, as determined by transaminase and creatinine measurements. Intranasal, but not intramuscular vaccination with SaAg led to a significant reduction in IL-10 production and was associated with increased level of IFN-γ and NO. SaAg intranasal vaccination was able to prime cellular and humoral immune responses and inducing a higher proliferation index and increased production of specific IgG1/IgG2, which contributed to decrease the bacterial load in both liver and the spleen and improve survival during sepsis. These findings present the first evidence of the effectiveness of whole Ag intranasal-based vaccine administration, which expands the vaccination possibilities against S. aureus infection. PMID:27091687

  2. Key regulators of sensitization and tolerance: GM-CSF, IL-10, TGF-β and the Notch signaling pathway in adjuvant-free experimental models of respiratory allergy.

    Guibas, George V; Makris, Michael; Papadopoulos, Nikolaos G

    2013-06-01

    Conventional experimental models of respiratory allergy have contributed greatly to our current knowledge of the pathophysiology of allergic airway diseases; nevertheless, they are contingent upon unnatural sensitization techniques, entailing adjuvant-aided intraperitoneal (i.p) administration of antigen. Currently, there is a growing appreciation of the impact of tolerance mechanics in the pathophysiology of respiratory allergy. Thus, inasmuch as adjuvants exert a robust tolerance-modifying action, a transition from the conventional method of experimental sensitization to one that is more naturally and clinically relevant becomes important. We therefore opted to survey the literature and identify agents that could interfere with sensitization mechanics following non-adjuvant-aided airway exposure of laboratory rodents to aeroallergen. GM-CSF was found to exert robust Th2-polarizing action in this setting. Conversely, IL-10 fulfilled an important, albeit not so clear-cut, tolerance-favoring role; TGF-β was also identified as a likely instigator of tolerogenesis. The role of Notch signaling in the sensitization versus tolerance dilemma appeared to be important but diverse. Collectively, these factors appeared to profoundly and diversely modulate the balance between tolerance and sensitization in naturally relevant experimental models of allergic airway disease. PMID:23768176

  3. Successful adjuvant-free vaccination of BALB/c mice with mutated amyloid β peptides

    Wahi Monika M

    2008-02-01

    Full Text Available Abstract Background A recent human clinical trial of an Alzheimer's disease (AD vaccine using amyloid beta (Aβ 1–42 plus QS-21 adjuvant produced some positive results, but was halted due to meningoencephalitis in some participants. The development of a vaccine with mutant Aβ peptides that avoids the use of an adjuvant may result in an effective and safer human vaccine. Results All peptides tested showed high antibody responses, were long-lasting, and demonstrated good memory response. Epitope mapping indicated that peptide mutation did not lead to epitope switching. Mutant peptides induced different inflammation responses as evidenced by cytokine profiles. Ig isotyping indicated that adjuvant-free vaccination with peptides drove an adequate Th2 response. All anti-sera from vaccinated mice cross-reacted with human Aβ in APP/PS1 transgenic mouse brain tissue. Conclusion Our study demonstrated that an adjuvant-free vaccine with different Aβ peptides can be an effective and safe vaccination approach against AD. This study represents the first report of adjuvant-free vaccines utilizing Aβ peptides carrying diverse mutations in the T-cell epitope. These largely positive results provide encouragement for the future of the development of human vaccinations for AD.

  4. Cyclosporine Inhibits Apoptosis in Experimental Murine Xerophthalamia Conjunctival Epithelium

    SUN Jinghua; WANG Jingxin

    2006-01-01

    apoptosis and protect goblet cell against the loss in experimental murine xerophathala-mia. Inhibition of apoptosis appears to be a key mechanism responsible for the therapeutic effect of CsA on xerophthalamia.

  5. Nanogel antigenic protein-delivery system for adjuvant-free intranasal vaccines.

    Nochi, Tomonori; Yuki, Yoshikazu; Takahashi, Haruko; Sawada, Shin-ichi; Mejima, Mio; Kohda, Tomoko; Harada, Norihiro; Kong, Il Gyu; Sato, Ayuko; Kataoka, Nobuhiro; Tokuhara, Daisuke; Kurokawa, Shiho; Takahashi, Yuko; Tsukada, Hideo; Kozaki, Shunji; Akiyoshi, Kazunari; Kiyono, Hiroshi

    2010-07-01

    Nanotechnology is an innovative method of freely controlling nanometre-sized materials. Recent outbreaks of mucosal infectious diseases have increased the demands for development of mucosal vaccines because they induce both systemic and mucosal antigen-specific immune responses. Here we developed an intranasal vaccine-delivery system with a nanometre-sized hydrogel ('nanogel') consisting of a cationic type of cholesteryl-group-bearing pullulan (cCHP). A non-toxic subunit fragment of Clostridium botulinum type-A neurotoxin BoHc/A administered intranasally with cCHP nanogel (cCHP-BoHc/A) continuously adhered to the nasal epithelium and was effectively taken up by mucosal dendritic cells after its release from the cCHP nanogel. Vigorous botulinum-neurotoxin-A-neutralizing serum IgG and secretory IgA antibody responses were induced without co-administration of mucosal adjuvant. Importantly, intranasally administered cCHP-BoHc/A did not accumulate in the olfactory bulbs or brain. Moreover, intranasally immunized tetanus toxoid with cCHP nanogel induced strong tetanus-toxoid-specific systemic and mucosal immune responses. These results indicate that cCHP nanogel can be used as a universal protein-based antigen-delivery vehicle for adjuvant-free intranasal vaccination. PMID:20562880

  6. Ureaplasma urealyticum Causes Hyperammonemia in an Experimental Immunocompromised Murine Model

    Wang, Xiaohui; Karau, Melissa J.; Greenwood-Quaintance, Kerryl E.; Block, Darci R.; Mandrekar, Jayawant N.; Cunningham, Scott A.

    2016-01-01

    Hyperammonemia syndrome is an often fatal complication of lung transplantation which has been recently associated with Ureaplasma infection. It has not been definitely established that Ureaplasma species can cause hyperammonemia. We established a novel immunocompromised murine model of Ureaplasma urealyticum infection and used it to confirm that U. urealyticum can cause hyperammonemia. Male C3H mice were pharmacologically immunosuppressed with mycophenolate mofetil, tacrolimus and oral prednisone for seven days, and then challenged intratracheally (IT) and/or intraperitoneally (IP) with 107 CFU U. urealyticum over six days, while continuing immunosuppression. Spent U. urealyticum-free U9 broth was used as a negative control, with uninfected immunocompetent mice, uninfected immunosuppressed mice, and infected immunocompetent mice serving as additional controls. Plasma ammonia concentrations were compared using Wilcoxon ranks sum tests. Plasma ammonia concentrations of immunosuppressed mice challenged IT/IP with spent U9 broth (n = 14) (range 155–330 μmol/L) were similar to those of normal mice (n = 5), uninfected immunosuppressed mice (n = 5), and U. urealyticum IT/IP challenged immunocompetent mice (n = 5) [range 99–340 μmol/L, p = 0.60]. However, immunosuppressed mice challenged with U. urealyticum IT/IP (n = 20) or IP (n = 15) had higher plasma ammonia concentrations (range 225–945 μmol/L and 276–687 μmol/L, respectively) than those challenged IT/IP with spent U9 broth (p<0.001). U. urealyticum administered IT/IP or IP causes hyperammonemia in mice pharmacologically immunosuppressed with a regimen similar to that administered to lung transplant recipients. PMID:27537683

  7. Costimulatory signal blockade in murine relapsing experimental autoimmune encephalomyelitis

    Schaub, M; Issazadeh-Navikas, Shohreh; Stadlbauer, T H;

    1999-01-01

    Blockade of the CD28-B7 or CD40L-CD40 T cell costimulatory signals prevents induction of experimental autoimmune encephalomyelitis (EAE). However, the effect of simultaneous blockade of these signals in EAE is unknown. We show that administration of either MR1 (to block CD40L) or CTLA4Ig (to bloc...

  8. Immune response to controlled release of immunomodulating peptides in a murine experimental autoimmune encephalomyelitis (EAE) model

    Zhao, Hong; Kiptoo, Paul; Williams, Todd D.; Siahaan, Teruna J.; Topp, Elizabeth M.

    2009-01-01

    The effects of controlled release on immune response to an immunomodulating peptide were evaluated in a murine experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS). The peptide, Ac-PLP-BPI-NH2-2 (Ac-HSLGKWLGHPDKF-(AcpGAcpGAcp)2-ITDGEATDSG-NH2; Ac = acetyl, Acp = aminocaproic acid) was designed to suppress T-cell activation in response to PLP139–151, an antigenic peptide in MS. Poly-lactide-co-glycolide (PLGA) microparticles containing Ac-PLP-BPI-NH2-2 (8±4 μm, 1.4±...

  9. Effects of vitamin E on mitochondrial dysfunction and asthma features in an experimental allergic murine model

    Mabalirajan, Ulaganathan; Aich, Jyotirmoi; Leishangthem, Geeta Devi; Sharma, Surendra Kumar; Dinda, Amit Kumar; Ghosh, Balaram

    2009-01-01

    We showed recently that IL-4 causes mitochondrial dysfunction in allergic asthma. IL-4 is also known to induce 12/15-lipoxygenase (12/15-LOX), a potent candidate molecule in asthma. Because vitamin E (Vit-E) reduces IL-4 and inhibits 12/15-LOX in vitro, here we tested the hypothesis that Vit-E may be effective in restoring key mitochondrial dysfunctions, thus alleviating asthma features in an experimental allergic murine model. Ovalbumin (OVA)-sensitized and challenged male BALB/c mice showed...

  10. A comprehensive collection of experimentally validated primers for Polymerase Chain Reaction quantitation of murine transcript abundance

    Wang Xiaowei

    2008-12-01

    Full Text Available Abstract Background Quantitative polymerase chain reaction (QPCR is a widely applied analytical method for the accurate determination of transcript abundance. Primers for QPCR have been designed on a genomic scale but non-specific amplification of non-target genes has frequently been a problem. Although several online databases have been created for the storage and retrieval of experimentally validated primers, only a few thousand primer pairs are currently present in existing databases and the primers are not designed for use under a common PCR thermal profile. Results We previously reported the implementation of an algorithm to predict PCR primers for most known human and mouse genes. We now report the use of that resource to identify 17483 pairs of primers that have been experimentally verified to amplify unique sequences corresponding to distinct murine transcripts. The primer pairs have been validated by gel electrophoresis, DNA sequence analysis and thermal denaturation profile. In addition to the validation studies, we have determined the uniformity of amplification using the primers and the technical reproducibility of the QPCR reaction using the popular and inexpensive SYBR Green I detection method. Conclusion We have identified an experimentally validated collection of murine primer pairs for PCR and QPCR which can be used under a common PCR thermal profile, allowing the evaluation of transcript abundance of a large number of genes in parallel. This feature is increasingly attractive for confirming and/or making more precise data trends observed from experiments performed with DNA microarrays.

  11. Comparison of histopathology and PCR based assay for detection of experimentally induced toxoplasmosis in murine model

    Vikrant Sudan; A K Tewari; R Singh; Harkirat Singh

    2015-01-01

    Objective:To compare histopathology and PCR based detection in diagnosis of experimentally induced toxoplasmosis of RH human strain of the parasite in murine models. Methods:A comparison of histopathology and PCR based detection was done to diagnose experimentally induced toxoplasmosis in ten inbred swiss albino mice after intraperitoneal inoculation of 100 tachyzoites of laboratory mantained human RH strain of the parasite. Tissue samples from lung, liver, spleen, brain, heart and kidney were taken and processed for histopathological examination while all the samples also were subjected to PCR, using primers directed to the multicopy of SAG 3 gene, in dublicates. Results: Histopathology revealed presence of tachyzoites only in liver while along with lung, liver, spleen and brain tissue yielded desired positive PCR amplicons. Conclusions:The SAG 3 based PCR is able to diagnose toxoplasmosis in those tissues which are declared negative by histopathological assay.

  12. Comparison of histopathology and PCR based assay for detection of experimentally induced toxoplasmosis in murine model

    Vikrant; Sudan; A.K.Tewari; R; Singh; Harkirat; Singh

    2015-01-01

    Objective:To compare histopathology and PCR based detection in diagnosis of experimentally induced toxoplasmosis of RH human strain of the parasite in murine models.Methods:A comparison of histopathology and PCR based detection was done to diagnose experimentally induced toxoplasmosis in ten inbred swiss albino mice after intraperitoneal inoculation of 100 tachyzoites of laboratory mantained human RH strain of the parasite.Tissue samples from lung,liver,spleen,brain,heart and kidney were taken and processed for histopathological examination while all the samples also were subjected to PCR,using primers directed to the multicopy of SAG 3 gene,in dublicates.Results:Histopathology revealed presence of tachyzoites only in liver while along with lung,liver,spleen and brain tissue yielded desired positive PCR amplicons.Conclusions:The SAG 3 based PCR is able to diagnose toxoplasmosis in those tissues which are declared negative by histopathological assay.

  13. Induction of protection in murine experimental models against Trichinella spiralis: an up-to-date review.

    Ortega-Pierres, G; Vaquero-Vera, A; Fonseca-Liñán, R; Bermúdez-Cruz, R M; Argüello-García, R

    2015-09-01

    The parasitic nematode Trichinella spiralis, an aetiological agent of the disease known as trichinellosis, infects wild and domestic animals through contaminated pig meat, which is the major source for Trichinella transmission. Prevention of this disease by interrupting parasite transmission includes vaccine development for livestock; however, major challenges to this strategy are the complexity of the T. spiralis life cycle, diversity of stage-specific antigens, immune-evasion strategies and the modulatory effect of host responses. Different approaches have been taken to induce protective immune responses by T. spiralis immunogens. These include the use of whole extracts or excretory-secretory antigens, as well as recombinant proteins or synthesized epitopes, using murine experimental models for trichinellosis. Here these schemes are reviewed and discussed, and new proposals envisioned to block the zoonotic transmission of this parasite. PMID:25761655

  14. Experimental infection of Phlebotomus perniciosus by bioluminescent Leishmania infantum using murine model and artificial feeder

    Cannet, Arnaud; Akhoundi, Mohammad; Michel, Gregory; Marty, Pierre; Delaunay, Pascal

    2016-01-01

    Leishmaniasis is a vector-borne disease that is transmitted by sandflies and caused by obligate intracellular protozoa of the genus Leishmania. In the present study, we carried out a screening on the experimental infection of Phlebotomus pernioucus by bioluminescent Leishmania infantum using murine model and artificial feeder. We developed a real-time polymerase chain reaction (RT-PCR)-based method to determine individually the number of Leishmania promastigotes fed by infected flies. Among 1840 new emerged female sand flies, 428 were fed on the infected mice. After their death, they were analysed individually by RT-PCR. Our results demonstrated just a single Leishmania positive female at sixth day post meal. A total of 1070 female sand flies were exposed in contact with artificial feeder containing the human blood with two different quantities of Leishmania parasites: 2.106/mL and 1.107/mL. A blood meal including 1.107/mL LUC-promastigotes was proposed to 270 females and 75 (28%) flies were engorged. Among them, 44 (59%) were positive by RT-PCR analysis, with a relative average of 50551 Leishmania parasites. In case of blood feeding of females with 2.106/mL promastigotes, 57 out of 800 (7%) females succeed to feed from artificial feeder which 22 (39%) were positive with a relative average of 6487 parasites. PMID:27439032

  15. Experimental murine chromoblastomycosis obtained from Fonsecaea pedrosoi isolate cultured for a long periodt

    AP Machado

    2009-01-01

    Full Text Available The present study aimed to describe F. pedrosoi propagules capable of causing chronic murine disease. Several changes in F. pedrosoi hyphae were identified in fungal cells cultured for a long period. Optical microscopy found many rounded cells with double-rigid melanin-rich walls. Terminal and intercalary chlamydoconidia were also frequently observed. Analyses of images from transmission electron microscopy (TEM revealed several cells with walls composed of at least three layers and an outer layer enriched with melanin. Two groups of twenty BALB/c mice were subcutaneously infected in their footpads with F. pedrosoi cells at an inoculum concentration of approximately 1 x 10(4 cells/mL. In one group, long-term cultured F. pedrosoi cells were inoculated in one footpad, whereas in the other group, both footpads were infected. Active lesions were observed up to seven months post-infection, particularly in mice inoculated at two sites. After this period, animals were killed. Histological sections revealed characteristics bearing a strong resemblance to the human form of the disease such as tissue hyperplasia, granulomas with microabscesses and sclerotic cells. Based on this study, we identified fungal cells from old cultures capable of provoking chronic chromoblastomycosis under experimental conditions, especially when more than one site is infected.

  16. Lipid alterations in experimental murine colitis: role of ceramide and imipramine for matrix metalloproteinase-1 expression.

    Jessica Bauer

    Full Text Available BACKGROUND: Dietary lipids or pharmacologic modulation of lipid metabolism are potential therapeutic strategies in inflammatory bowel disease (IBD. Therefore, we analysed alterations of bioactive lipids in experimental models of colitis and examined the functional consequence of the second messenger ceramide in inflammatory pathways leading to tissue destruction. METHODOLOGY/PRINCIPAL FINDINGS: Chronic colitis was induced by dextran-sulphate-sodium (DSS or transfer of CD4(+CD62L(+ cells into RAG1(-/--mice. Lipid content of isolated murine intestinal epithelial cells (IEC was analysed by tandem mass spectrometry. Concentrations of MMP-1 in supernatants of Caco-2-IEC and human intestinal fibroblasts from patients with ulcerative colitis were determined by ELISA. Imipramine was used for pharmacologic inhibition of acid sphingomyelinase (ASM. Ceramide increased by 71% in chronic DSS-induced colitis and by 159% in the transfer model of colitis. Lysophosphatidylcholine (LPC decreased by 22% in both models. No changes were detected for phosphatidylcholine. Generation of ceramide by exogenous SMase increased MMP-1-protein production of Caco-2-IEC up to 7-fold. Inhibition of ASM completely abolished the induction of MMP-1 by TNF or IL-1beta in Caco-2-IEC and human intestinal fibroblasts. CONCLUSIONS/SIGNIFICANCE: Mucosal inflammation leads to accumulation of ceramide and decrease of LPC in the intestinal epithelium. One aspect of ceramide generation is an increase of MMP-1. Induction of MMP-1 by TNF or IL-1beta is completely blocked by inhibition of ASM with imipramine. Therefore, inhibition of ASM may offer a treatment strategy to reduce MMP-1 expression and tissue destruction in inflammatory conditions.

  17. Cellular basis of the genetic susceptibility of murine experimental allergic encephalomyelitis

    Murine experimental allergic encephalomyelitis (EAE) is an induced autoimmune disease that resembles human multiple sclerosis. The authors have investigated the cellular basis of the genetic predisposition and resistance of inbred strains of mice to EAE using an adoptive transfer system between two H-2 compatible, Thy 1 antigen disparate strains of mice. Genetically EAE susceptible SJL/J strain mice (H-2/sup s/, Thy 1.2) and resistant B10.S Thy 1.1 (H-2/sub s/, Thy 1.1) strain mice were lethally irradiated (700R) and reconstituted with 5-10 x 106 bone marrow cells from either SJL/J or congenic B10.S (Thy 1.1 or Thy 1.2) donors. After 30-45 days, more than 95% of the thymocytes and 75% of the peripheral T cells in the chimeras were of donor origin. These lymphohemopoietic chimeras were then sensitized in their hind footpads with porcine myelin basic protein in complete Freund's adjuvant containing M. tuberculosis H37RA, followed at 24 and 72 hours by i.v. injection of B. pertussis. Clinical signs of EAE developed in unirradiated SJL/J, but not B10.S, controls, and in irradiated B10.S and SJL/J recipients of SJL/J, but not B10.S, bone marrow. These results indicate that bone marrow cells can transfer the predisposition to EAE from genetically susceptible to genetically resistant mouse strains. The cellular component in the bone marrow that is responsible for the transfer of the genetic susceptibility to EAE is under investigation

  18. Protein tyrosine phosphatase 1B deficiency ameliorates murine experimental colitis via the expansion of myeloid-derived suppressor cells.

    Jing Zhang

    Full Text Available Protein tyrosine phosphatase 1B (PTP1B is a key molecule in modulating low-degree inflammatory conditions such as diabetes. The role of PTP1B in other chronic inflammations, however, remains unknown. Here, we report that PTP1B deficiency ameliorates Dextran Sulfate Sodium (DSS-induced murine experimental colitis via expanding CD11b(+Gr-1(+ myeloid-derived suppressor cells (MDSCs. Employing DSS-induced murine experimental colitis as inflammatory animal model, we found that, compared with wild-type littermates, PTP1B-null mice demonstrated greater resistance to DSS-induced colitis, as reflected by slower weight-loss, greater survival rates and decreased PMN and macrophage infiltration into the colon. The evidence collectively also demonstrated that the resistance of PTP1B-null mice to DSS-induced colitis is based on the expansion of MDSCs. First, PTP1B-null mice exhibited a greater frequency of MDSCs in the bone marrow (BM, peripheral blood and spleen when compared with wild-type littermates. Second, PTP1B levels in BM leukocytes were significantly decreased after cells were induced into MDSCs by IL-6 and GM-CSF, and the MDSC induction occurred more rapidly in PTP1B-null mice than in wild-type littermates, suggesting PTP1B as a negative regulator of MDSCs. Third, the adoptive transfer of MDSCs into mice with DSS-colitis significantly attenuated colitis, which accompanies with a decreased serum IL-17 level. Finally, PTP1B deficiency increased the frequency of MDSCs from BM cells likely through enhancing the activities of signal transducer and activator of transcription 3 (STAT3 and Janus kinase 2 (JAK2. In conclusion, our study provides the first evidences that PTP1B deficiency ameliorates murine experimental colitis via expanding MDSCs.

  19. Efficacy of Lysophosphatidylcholine in Combination with Antimicrobial Agents against Acinetobacter baumannii in Experimental Murine Peritoneal Sepsis and Pneumonia Models.

    Parra Millán, R; Jiménez Mejías, M E; Sánchez Encinales, V; Ayerbe Algaba, R; Gutiérrez Valencia, A; Pachón Ibáñez, M E; Díaz, C; Pérez Del Palacio, J; López Cortés, L F; Pachón, J; Smani, Y

    2016-08-01

    Immune response stimulation to prevent infection progression may be an adjuvant to antimicrobial treatment. Lysophosphatidylcholine (LPC) is an immunomodulator involved in immune cell recruitment and activation. In this study, we aimed to evaluate the efficacy of LPC in combination with colistin, tigecycline, or imipenem in experimental murine models of peritoneal sepsis and pneumonia. We used Acinetobacter baumannii strain Ab9, which is susceptible to colistin, tigecycline, and imipenem, and multidrug-resistant strain Ab186, which is susceptible to colistin and resistant to tigecycline and imipenem. Pharmacokinetic and pharmacodynamic parameters for colistin, tigecycline, and imipenem and the 100% minimal lethal dose (MLD100) were determined for both strains. The therapeutic efficacies of LPC, colistin (60 mg/kg of body weight/day), tigecycline (10 mg/kg/day), and imipenem (180 mg/kg/day), alone or in combination, were assessed against Ab9 and Ab186 at the MLD100 in murine peritoneal sepsis and pneumonia models. The levels of pro- and anti-inflammatory cytokines, i.e., tumor necrosis factor alpha (TNF-α) and interleukin-10 (IL-10), were determined by enzyme-linked immunosorbent assay (ELISA) for the same experimental models after inoculating mice with the MLD of both strains. LPC in combination with colistin, tigecycline, or imipenem markedly enhanced the bacterial clearance of Ab9 and Ab186 from the spleen and lungs and reduced bacteremia and mouse mortality rates (P colistin, tigecycline, and imipenem monotherapies. Moreover, at 4 h post-bacterial infection, Ab9 induced higher TNF-α and lower IL-10 levels than those with Ab186 (4 μg/ml versus 3 μg/ml [P colistin, tigecycline, or imipenem modestly reduced the severity of infection by A. baumannii strains with different resistance phenotypes compared to LPC monotherapy in both experimental models. PMID:27161639

  20. Pathobiology of human RH strain induced experimental toxoplasmosis in murine model.

    Sudan, Vikrant; Tewari, A K; Singh, Harkirat; Singh, R

    2016-09-01

    Of late, toxoplasmosis has gained immense importance as an opportunist parasite in immunocompromised patients. In immunocompromised subjects, the disease is supposed to occur in acute form and causes acute toxoplasmic encephalitis. However, the exact pathogenesis of other vital organs, particularly in acute form of infection, is still a matter of debate. Therefore, an attempt was made to study the pathogenesis of acute form of toxoplasmosis using cryopreserved human RH strain of the parasite in murine models. For this, 100 tachyzoites were given to individual mice and upon the setup of acute form of infection, the mice were euthanized and the organs were processed for histopathology. Histopathology revealed tachyzoites in liver only while severe necrosis due to multiplication of tachyzoites were visible in liver, spleen, lungs and brain. Kidneys and heart appeared more or less normal. Finally, the pathology of disease in these organs is described in detail. The present research has generated some vital information regarding necrotic changes in tissues due to acute toxoplasmosis and will defiantly help the researchers in the better understanding of disease particularly in humans and putting up of suitable treatment regime for human subjects infected with acute toxoplasmosis. PMID:27605794

  1. Evaluation of therapeutic potential of nanosilver particles synthesised using aloin in experimental murine mastitis model.

    Chaitanya Kumar, Thota Venkata; Muralidhar, Yegireddy; Prasad, Pagadala Eswara; Prasad, Tollamadugu Naga Venkata Krishna Vara; Alpha Raj, Mekapogu

    2013-09-01

    Nanobiotechnology is an emerging biological branch of nanotechnology. Application of nanoparticles with specific size and shape in biology has already shown unforeseen and interesting results. A study was conducted to evaluate the therapeutic potential of phytogenically derived aloin mediated nanosilver particles (AAgNPs), prepared by reduction of silver nitrate with aloin, in Staphylococcus aureus induced murine mastitis. A total of 40 female mice were divided into five groups of eight animals each. Group I served as lactating control, groups II-V were inoculated with 20 μl of 24 h broth culture of S. aureus containing 4.0 × 105 cfu/quarter under ketamine anaesthesia. After 6 h post inoculation, groups III and IV received 20 μl of aloin nanosilver (AAgNPs) through intramammary and intraperitoneal routes, respectively. Group V received antibiotic cefepime at 1 mg/kg body weight through the intra-peritoneal route. After 18 h post-treatment, serum C reactive protein, weights of mammary glands, mammary gland bacterial load, thiobarbituric acid reactive substances content, reduced glutathione content, superoxide dismutase activity and catalase activity and histopathology were determined. The compound showed a minimum inhibitory concentration of 21.8 ng/ml against S. aureus. Significant reduction (98%) in poly-morpho nuclear cell infiltration was observed with AAgNPs than antibiotic (50%). PMID:24028805

  2. Ultrastructural study on experimental infection of rotavirus in a murine heterologous model

    Selma Majerowicz

    1994-09-01

    Full Text Available Viral replication, histopathological and ultrastructural changes were observed for a period of nine days in the small intestine of suckling mice infected with a simian rotavirus (SA11. Samples taken from duodenum, jejunun and ileum were prepared for light microscopy, transmission and scanning electron microscopy analysis. Histopathologic effect could be detected within 8 hr post-infection, when only a few altered cells were observed. Damage was extensive after 16 hr post-infection, showing swollen enterocytes and reduced and irregularly oriented microvilli at intestinal villi tips. Virus particles were detected at 16 and 48 hr post-infection, budding from the viroplasm into the rough endoplasmic reticulum cisternae in ileum enterocytes. Clear evidence of viral replication, observed by electron microscopy was not described before in heterologous murine models. Regeneration of the intestinal villi began at the third day post-infection. Despite some differences observed in clinical symptoms and microscopic analysis of homologous and heterologous rotavirus infections, we concluded that mechanisms of heterologous rotavirus infection in mice follow similar patterns to those observed in the homologous models.

  3. Prevention of murine experimental autoimmune orchitis by recombinant human interleukin-6

    Li, Lu; Itoh, Masahiro; Ablake, Maila;

    2002-01-01

    We studied the effect of exogenously administered recombinant human interleukin (IL)-6 on the development of experimental autoimmune orchitis (EAO) in C3H/Hej mice. IL-6 significantly reduced histological signs of EAO and appearance of delayed type hypersensitivity against the immunizing testicular...

  4. Utility of the microculture method in non-invasive samples obtained from an experimental murine model with asymptomatic leishmaniasis.

    Allahverdiyev, Adil M; Bagirova, Malahat; Cakir-Koc, Rabia; Elcicek, Serhat; Oztel, Olga Nehir; Canim-Ates, Sezen; Abamor, Emrah Sefik; Yesilkir-Baydar, Serap

    2012-07-01

    The sensitivity of diagnostic methods for visceral leishmaniasis (VL) decreases because of the low number of parasites and antibody amounts in asymptomatic healthy donors who are not suitable for invasive sample acquisition procedures. Therefore, new studies are urgently needed to improve the sensitivity and specificity of the diagnostic approaches in non-invasive samples. In this study, the sensitivity of the microculture method (MCM) was compared with polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and immunofluorescent antibody test (IFAT) methods in an experimental murine model with asymptomatic leishmaniasis. Results showed that the percent of positive samples in ELISA, IFAT, and peripheral blood (PB) -PCR tests were 17.64%, 8.82%, and 5.88%, respectively, whereas 100% positive results were obtained with MCM and MCM-PCR methods. Thus, this study, for the first time, showed that MCM is more sensitive, specific, and economic than other methods, and the sensitivity of PCR that was performed to samples obtained from MCM was higher than sensitivity of the PCR method sampled by PB. PMID:22764296

  5. Sepsis otopathy: experimental sepsis leads to significant hearing impairment due to apoptosis and glutamate excitotoxicity in murine cochlea

    Joachim Schmutzhard

    2013-05-01

    Hearing loss is frequent in intensive care patients and can be due to several causes. However, sepsis has not been examined as a possible cause. The aim of this study is to assess the influence of experimental sepsis on hearing thresholds and to evaluate pathological changes in the cochlea. The cecal ligation puncture technique was used to induce sepsis in 18 mice. Results were compared with those from 13 sham-operated and 13 untreated control mice. The hearing thresholds of the animals were evaluated with auditory evoked brainstem responses prior to the induction of sepsis and again at the peak of the disease. Immediately after the second measurement, the mice were sacrificed and the inner ears harvested and prepared for further evaluation. The cochleae were examined with light microscopy, electron microscopy and immunohistochemistry for Bax, cleaved caspase-3 and Bcl-2. The mice with sepsis showed a significant hearing loss but not the control groups. Induction of apoptosis could be shown in the supporting cells of the organ of Corti. Furthermore, excitotoxicity could be shown at the basal pole of the inner hair cells. In this murine model, sepsis leads to significant hearing impairment. The physiological alteration could be linked to apoptosis in the supporting cells of the organ of Corti and to a disturbance of the synapses of the inner hair cells.

  6. Assessment and in vivo scoring of murine experimental autoimmune uveoretinitis using optical coherence tomography

    Chu, C J; Herrmann, P.; Carvalho, L. S.; Liyanage, S. E.; Bainbridge, J. W.; Ali, R. R.; Dick, A. D.; Luhmann, U. F.

    2013-01-01

    Despite advances in clinical imaging and grading our understanding of retinal immune responses and their morphological correlates in experimental autoimmune uveoretinitis (EAU), has been hindered by the requirement for post-mortem histology. To date, monitoring changes occurring during EAU disease progression and evaluating the effect of therapeutic intervention in real time has not been possible. We wanted to establish whether optical coherence tomography (OCT) could detect intraretinal chan...

  7. Prevention of murine experimental autoimmune orchitis by recombinant human interleukin-6

    Li, Lu; Itoh, Masahiro; Ablake, Maila;

    2002-01-01

    We studied the effect of exogenously administered recombinant human interleukin (IL)-6 on the development of experimental autoimmune orchitis (EAO) in C3H/Hej mice. IL-6 significantly reduced histological signs of EAO and appearance of delayed type hypersensitivity against the immunizing testicular...... germinal cells. The effect was seen even though the cytokine was administered for only 6 consecutive days and 2 weeks after immunization....

  8. Effects of ionizing radiation on bone cell differentiation in an experimental murine bone cell model

    Baumstark-Khan, Christa; Lau, Patrick; Hellweg, Christine; Reitz, Guenther

    During long-term space travel astronauts are exposed to a complex mixture of different radiation types under conditions of dramatically reduced weight-bearing activity. It has been validated that astronauts loose a considerable amount of bone mass at a rate up to one to two percent each month in space. Therapeutic doses of ionizing radiation cause bone damage and increase fracture risks after treatment for head-and-neck cancer and in pelvic irradiation. For low radiation doses, the possibility of a disturbed healing potential of bone was described. Radiation induced damage has been discussed to inflict mainly on immature and healing bone. Little is known about radiation effects on bone remodelling and even less on the combined action of microgravity and radiation. Bone remodelling is a life-long process performed by balanced action of cells from the osteoblast and osteoclast lineages. While osteoblasts differentiate either into bone-lining cells or into osteocytes and play a crucial role in bone matrix synthesis, osteoclasts are responsible for bone resorption. We hypothesize that the balance between bone matrix assembly by osteocytes and bone degradation by osteoclasts is modulated by microgravity as well as by ionizing radiation. To address this, a cell model consisting of murine cell lines with the potential to differentiate into bone-forming osteoblasts (OCT-1, MC3T3-E1 S24, and MC3T3-E1 S4) was used for studying radiation response after exposure to simulated components of cosmic radiation. Cells were exposed to graded doses of 150 kV X-rays, α particles (0.525 MeV/u, 160 keV/µm; PTB, Braunschweig, Germany) and accelerated heavy ions (75 MeV/u carbon, 29 keV/µm; 95 MeV/u argon, 230 keV/µm; GANIL, Caen, France). Cell survival was measured as colony forming ability; cell cycle progression was analyzed via fluorescence-activated cell scanning (FACS) by measurement of the content of propidium iodide-stained DNA, DNA damage was visualized by γH2AX

  9. Effect of piroxicam, metamizol, and S-adenosylmethionine in a murine model of experimental trichomoniasis

    Nogal-Ruiz J.J.

    2005-03-01

    Full Text Available Biological effects of piroxicam, metamizol, and S-adenosylmethionine (S-AMET have been tested in NMRI mice infected intraperitoneally with Trichomonas vaginalis. An intraperitoneal treatment during ten preinfection days with piroxicam (10 mg/Kg/day, or metamizol (275 mg/Kg/day, but not with S-AMET (17 mg/Kg/day induced a significant decrease of abdominal lesions and mortality, assessed by means of a pathogenicity index. The trichomonicidal activity of piroxicam, metamizol, and S-AMET was tested in vitro at the concentration of 300 μM, but found ineffective. These assays have shown the usefulness of the experimental trichomoniasis model for the study of the immunomodulating activity of synthetic drugs.

  10. Genetic deletion of dectin-1 does not affect the course of murine experimental colitis

    Heinsbroek Sigrid EM

    2012-04-01

    Full Text Available Abstract Background It is believed that inflammatory bowel diseases (IBD result from an imbalance in the intestinal immune response towards the luminal microbiome. Dectin-1 is a widely expressed pattern recognition receptor that recognizes fungi and upon recognition it mediates cytokine responses and skewing of the adaptive immune system. Hence, dectin-1 may be involved in the pathogenesis of IBD. Methods We assessed the responses of dectin-1 deficient macrophages to the intestinal microbiota and determined the course of acute DSS and chronic Helicobacter hepaticus induced colitis in dectin-1 deficient mice. Results We show that the mouse intestinal microbiota contains fungi and the cytokine responses towards this microbiota were significantly reduced in dectin-1 deficient macrophages. However, in two different colitis models no significant differences in the course of inflammation were found in dectin-1 deficient mice compared to wild type mice. Conclusions Together our data suggest that, although at the immune cell level there is a difference in response towards the intestinal flora in dectin-1 deficient macrophages, during intestinal inflammation this response seems to be redundant since dectin-1 deficiency in mice does not affect intestinal inflammation in experimental colitis.

  11. Systemic Inflammatory Effects of Traumatic Brain Injury, Femur Fracture, and Shock: An Experimental Murine Polytrauma Model

    C. Probst

    2012-01-01

    Full Text Available Objective. Despite broad research in neurotrauma and shock, little is known on systemic inflammatory effects of the clinically most relevant combined polytrauma. Experimental investigation in an animal model may provide relevant insight for therapeutic strategies. We describe the effects of a combined injury with respect to lymphocyte population and cytokine activation. Methods. 45 male C57BL/6J mice (mean weight 27 g were anesthetized with ketamine/xylazine. Animals were subjected to a weight drop closed traumatic brain injury (WD-TBI, a femoral fracture and hemorrhagic shock (FX-SH. Animals were subdivided into WD-TBI, FX-SH and combined trauma (CO-TX groups. Subjects were sacrificed at 96 h. Blood was analysed for cytokines and by flow cytometry for lymphocyte populations. Results. Mortality was 8%, 13% and 47% for FX-SH, WD-TBI and CO-TX groups (P<0.05. TNFα (11/13/139 for FX-SH/WD-TBI/CO-TX; P<0.05, CCL2 (78/96/227; P<0.05 and IL-6 (16/48/281; P=0.05 showed significant increases in the CO-TX group. Lymphocyte populations results for FX-SH, WD-TBI and CO-TX were: CD-4 (31/21/22; P= n.s., CD-8 (7/28/34, P<0.05, CD-4-CD-8 (11/12/18; P= n.s., CD-56 (36/7/8; P<0.05. Conclusion. This study shows that a combination of closed TBI and femur-fracture/ shock results in an increase of the humoral inflammation. More attention to combined injury models in inflammation research is indicated.

  12. An experimental study of artificial murine bladder reflex arc established by abdominal reflex

    WANG Jin-wu; ZHAO Yu-wu; HOU Chun-lin; NI Wei-feng; RUI Bi-yu; GUO Shang-chun; ZHENG Xian-you; DAI Ke-rong

    2011-01-01

    Background The neurogenic bladder dysfunction caused by spinal cord injury is difficult to treat clinically. The aim of this research was to establish an artificial bladder reflex arc in rats through abdominal reflex pathway above the level of spinal cord injury, reinnervate the neurogenic bladder and restore bladder micturition.Methods The outcome was achieved by intradural microanastomosis of the right T13 ventral root to S2 ventral root with autogenous nerve grafting, leaving the right T13 dorsal root intact. Long-term function of the reflex arc was assessed from nerve electrophysiological data and intravesical pressure tests during 8 months postoperation. Horseradish peroxidase (HRP) tracing was performed to observe the effectiveness of the artificial reflex.Results Single stimulus (3 mA, 0.3 ms pulses, 20 Hz, 5-second duration) on the right T13 dorsal root resulted in evoked action potentials, raised intravesical pressures and bladder smooth muscle, compound action potential recorded from the right vesical plexus before and after the spinal cord transaction injury between L5 and S4 segmental in 12 Sprague-Dawley rats. There were HRP labelled cells in T13 ventral horn on the experimental side and in the intermediolateral nucleus on both sides of the L6-S4 segments after HRP injection. There was no HRP labelled cell in T13 ventral horn on the control side.Conclusion Using the surviving somatic reflex above the level of spinal cord injury to reconstruct the bladder autonomous reflex arc by intradural microanastomosis of ventral root with a segment of autologous nerve grafting is practical in rats and may have clinical applications for humans.

  13. Macrophage Phenotype in the Ocular Surface of Experimental Murine Dry Eye Disease.

    You, In-Cheon; Coursey, Terry G; Bian, Fang; Barbosa, Flavia L; de Paiva, Cintia S; Pflugfelder, Stephen C

    2015-08-01

    To evaluate the phenotype of macrophages in the cornea and conjunctiva of C57BL/6 mice with induced experimental dry eye. C57BL/6 mice exposed to desiccating stress (DS) were evaluated at 1, 5, and 10 days and C57BL/6 mice maintained in non-stressed environment were used as controls. Whole eyes and adnexa were excised for histology or used for gene expression analysis. Location and phenotype of macrophages infiltrating the cornea and conjunctiva was evaluated by immunofluorescence analysis. Quantitative polymerase chain reaction evaluated macrophage markers and T cell-related and inflammatory cytokine expression in cornea and conjunctiva. Immunofluorescence staining demonstrated that macrophages reside in the conjunctiva of control and dry eye mice and their number did not change with DS. Real-time RT-PCR demonstrated that the level of M1 macrophage marker, iNOS, increased prominently in the conjunctiva at DS 10 days. In contrast, there was a non-significant decrease of the M2 marker Arg1 with DS. The levels of inflammatory cytokine, IL-12a mRNA transcript in the conjunctiva increased significantly at DS1 and decreased at DS5, while levels of IL-18 were significantly increased at DS 10. Macrophages reside in the ocular surface tissues of C57BL/6 mice. Although the number of macrophages in the conjunctiva does not change, evidence of inflammatory M1 activation after desiccating stress was observed. Better understanding of phagocyte diversity and activation in dry eye disease provide a basis for the development of phagocyte-targeted therapeutic strategies. PMID:25772203

  14. Estimating biologically relevant parameters under uncertainty for experimental within-host murine West Nile virus infection.

    Banerjee, Soumya; Guedj, Jeremie; Ribeiro, Ruy M; Moses, Melanie; Perelson, Alan S

    2016-04-01

    West Nile virus (WNV) is an emerging pathogen that has decimated bird populations and caused severe outbreaks of viral encephalitis in humans. Currently, little is known about the within-host viral kinetics of WNV during infection. We developed mathematical models to describe viral replication, spread and host immune response in wild-type and immunocompromised mice. Our approach fits a target cell-limited model to viremia data from immunocompromised knockout mice and an adaptive immune response model to data from wild-type mice. Using this approach, we first estimate parameters governing viral production and viral spread in the host using simple models without immune responses. We then use these parameters in a more complex immune response model to characterize the dynamics of the humoral immune response. Despite substantial uncertainty in input parameters, our analysis generates relatively precise estimates of important viral characteristics that are composed of nonlinear combinations of model parameters: we estimate the mean within-host basic reproductive number,R0, to be 2.3 (95% of values in the range 1.7-2.9); the mean infectious virion burst size to be 2.9 plaque-forming units (95% of values in the range 1.7-4.7); and the average number of cells infected per infectious virion to be between 0.3 and 0.99. Our analysis gives mechanistic insights into the dynamics of WNV infection and produces estimates of viral characteristics that are difficult to measure experimentally. These models are a first step towards a quantitative understanding of the timing and effectiveness of the humoral immune response in reducing host viremia and consequently the epidemic spread of WNV. PMID:27075003

  15. Experimental parameters differentially affect the humoral response of the cholera-toxin-based murine model of food allergy

    Kroghsbo, S.; Christensen, Hanne Risager; Frøkiær, Hanne

    2003-01-01

    Background: Recent studies have developed a murine model of IgE-mediated food allergy based on oral coadministration of antigen and cholera toxin (CT) to establish a maximal response for studying immunopathogenic mechanisms and immunotherapeutic strategies. However, for studying subtle...... interested in characterizing the individual effects of the parameters in the CT-based model: CT dose, antigen type and dose, and number of immunizations. Methods: BALB/c mice were orally sensitized weekly for 3 or 7 weeks with graded doses of CT and various food antigens (soy-trypsin inhibitor, ovalbumin or...... antibody response depended on the type of antigen and number of immunizations. Conclusions: The critical parameters of the CT-based murine allergy model differentially control the intensity and kinetics of the developing immune response. Adjustment of these parameters could be a key tool for tailoring the...

  16. Experimental parameters differentially affect the humoral response of the cholera-toxin-based murine model of food allergy

    Kroghsbo, S.; Christensen, Hanne Risager; Frøkiær, Hanne

    2003-01-01

    Background: Recent studies have developed a murine model of IgE-mediated food allergy based on oral coadministration of antigen and cholera toxin (CT) to establish a maximal response for studying immunopathogenic mechanisms and immunotherapeutic strategies. However, for studying subtle immunomodu......Background: Recent studies have developed a murine model of IgE-mediated food allergy based on oral coadministration of antigen and cholera toxin (CT) to establish a maximal response for studying immunopathogenic mechanisms and immunotherapeutic strategies. However, for studying subtle...... interested in characterizing the individual effects of the parameters in the CT-based model: CT dose, antigen type and dose, and number of immunizations. Methods: BALB/c mice were orally sensitized weekly for 3 or 7 weeks with graded doses of CT and various food antigens (soy-trypsin inhibitor, ovalbumin or...

  17. Immunogenic multistage recombinant protein vaccine confers partial protection against experimental toxoplasmosis mimicking natural infection in murine model

    Yaprak Gedik

    2016-01-01

    To generate a protective vaccine against toxoplasmosis, multistage vaccines and usage of challenging models mimicking natural route of infection are critical cornerstones. In this study, we generated a BAG1 and GRA1 multistage vaccine that induced strong immune response in which the protection was not at anticipated level. In addition, the murine model was orally challenged with tissue cysts to mimic natural route of infection.

  18. Expression and Significance of NF-κB, IL-1β and COX-2 in the Murine Model of Estrogen-dependent Experimental Vulvovaginal Candidiasis

    Xue-rong CHEN; Ya-li LIU; Dun-zhen XIAO; Jun GAO

    2007-01-01

    Objective To investigate the possible role of estrogen in the pathogenesis of vulvovaginal candidiasis(VVC).Methods Estrogen-dependent experimental murine model of C. albicans vaginal infection was established by injecting subcutaneously with estradiol benzoate and then 5 × 106 stationary-phase C. albicans blastoconidia was inoculated intravaginally to mice (group EI),and other 3 groups were set up: estrogen-treated but not infected (group E) ;estrogen-untreated but infected (group Ⅰ);normal control (group C).The dynamic change of colony-forming unit (CFU) of cervivovaginal lavage fluid was observed. Vaginal tissues at different time points (d 2,d 4,d 7 and d 14) after inoculation of C.albicans were obtained.In situ hybridization staining was used to detect expression of on d 4 and d 7 (P<0.01).Conclusions In the murine model of estrogen-dependent experimental VVC,estrogen promotes the infection establishment by up-regulating expression of CO X-2 via activating NF-κB signal pathway,and the high expression of COX-2 promoted by the interaction of IL-1β and NF-κB after infection formation was involved in persistence of infection.

  19. Interleukin-10 Enhances the Therapeutic Effectiveness of a Recombinant Poxvirus-Based Vaccine in an Experimental Murine Tumor Model

    Kaufman, Howard L.; Rao, Jay B.; Irivine, Kari R.; Bronte, Vincenzo; Rosenberg, Steven A.; Restifo, Nicholas P

    1999-01-01

    Interleukin-10 (IL-10) has a wide range of in vivo biological activities and is a key regulatory cytokine of immune-mediated inflammation. The authors found that murine IL-10 given 12 hours after a recombinant vaccinia virus (rVV) containing the LacZ gene significantly enhanced the treatment of mice bearing 3-day-old pulmonary metastases expressing β-galactosidase. Because IL-10 has been shown to inhibit the functions of key elements of both innate and acquired immune responses, the authors h...

  20. Rosmarinus officinalis L. extract ameliorates intestinal inflammation through MAPKs/NF-κB signaling in a murine model of acute experimental colitis.

    Medicherla, Kanakaraju; Ketkar, Avanee; Sahu, Bidya Dhar; Sudhakar, Godi; Sistla, Ramakrishna

    2016-07-13

    We investigated the anti-inflammatory and anti-colitis effects of Rosmarinus officinalis L. extract (RE) by using both in vitro LPS-activated mouse RAW 264.7 macrophages and in vivo dextran sulfate sodium (DSS)-induced experimental murine colitis and suggested the underlying possible mechanisms. Liquid Chromatography-Mass Spectrometry (LC-MS) analysis was performed to identify the major components present in the RE. The clinical signs, biochemistry, immunoblot, ELISA and histology in colon tissues were assessed in order to elucidate the beneficial effect of RE. RE suppressed the LPS-induced pro-inflammatory cytokine production and the expressions of inflammatory proteins in macrophages. Administration of RE (50 and 100 mg kg(-1)) also significantly reduced the severity of DSS-induced murine colitis, as assessed by the clinical symptoms, colon length and histology. RE administration prevented the DSS-induced activation of p38, ERK and JNK MAPKs, attenuated IκBα phosphorylation and subsequent nuclear translocation and DNA binding of NF-κB (p65). RE also suppressed the COX-2 and iNOS expressions, decreased the levels of TNF-α and IL-6 cytokines and the myeloperoxidase activity in the colon tissue. Histological observation revealed that RE administration alleviated mucosal damage and inflammatory cell infiltration induced by DSS in the colon tissue. Hence, RE could be used as a new preventive and therapeutic food ingredient or as a dietary supplement for inflammatory bowel disease. PMID:27349640

  1. Time course of reoxygenation in experimental murine tumors after carbon-beam and X-ray irradiation

    We compared the tumor reoxygenation patterns in three different murine tumor cell lines after X-irradiation with those after carbon-beam irradiation using a heavy-ion medical accelerator (HIMAC) system. The tumors of the cell lines SCCVII, SCCVII-variant-1 and EMT6 on the hind legs of mice received local priming irradiation with a carbon-beam (8 Gy, 73 keV/μm in LET, 290 MeV/u, 6 cm SOBP) or X-rays (13 Gy, 250 kVp). After various intervals, the mice were given whole-body test irradiation (16 Gy, 250 kVp X-ray) either in air or after they were killed. The hypoxic fractions were estimated as the proportions of the surviving fractions of the tumors in killed mice to those in air-breathing mice. In the SCCVII tumors, the hypoxic fractions at 0.5 h were 50% and 21% (p<0.05) after the priming x-irradiation and carbon-beam irradiation, respectively. In the SCCVII-variant-1 tumors, the hypoxic fractions were 85% and 82% at 0.5 h, 84% and 20% at 12 h (p<0.01), and 21% and 31% at 24 h after X-ray and after carbon-beam irradiation, respectively. In the EMT6 tumors, the reoxygenation patterns after X-irradiation and carbon-beam irradiation were quite similar. We concluded that the reoxygenation pattern differed among the three tumor cell lines, and that reoxygenation tended to occur more rapidly after carbon-beam irradiation than after X-irradiation for SCCVII and SCCVII-variant-1 tumors. (author)

  2. Eosinophilic esophagitis: published evidences for disease subtypes, indications for patient subpopulations, and how to translate patient observations to murine experimental models.

    Mudde, Anne C A; Lexmond, Willem S; Blumberg, Richard S; Nurko, Samuel; Fiebiger, Edda

    2016-01-01

    Eosinophilic esophagitis (EoE) is a chronic inflammatory disorder of the esophagus and commonly classified as a Th2-type allergy. Major advances in our understanding of the EoE pathophysiology have recently been made, but clinicians struggle with highly unpredictable therapy responses indicative of phenotypic diversity within the patient population. Here, we summarize evidences for the existence of EoE subpopulations based on diverse inflammatory characteristics of the esophageal tissue in EoE. Additionally, clinical characteristics of EoE patients support the concept of disease subtypes. We conclude that clinical and experimental evidences indicate that EoE is an umbrella term for conditions that are unified by esophageal eosinophilia but that several disease subgroups with various inflammatory esophageal patterns and/or different clinical features exist. We further discuss strategies to study the pathophysiologic differences as observed in EoE patients in murine experimental EoE. Going forward, models of EoE that faithfully mimic EoE subentities as defined in humans will be essential because mechanistic studies on triggers which regulate the onset of diverse EoE subpopulations are not feasible in patients. Understanding how and why different EoE phenotypes develop will be a first and fundamental step to establish strategies that integrate individual variations of the EoE pathology into personalized therapy. PMID:27458501

  3. Does Carica papaya leaf-extract increase the platelet count? An experimental study in a murine model

    Susiji Wickramasinghe; Roshitha Nilmini Waduge

    2013-01-01

    Objective:To investigate the potential role of fresh Carica papaya (C. papaya) leaf extract on haematological and biochemical parameters and toxicological changes in a murine model. Methods: In total 36 mice were used for the trial. Fresh C. papaya leaf extract [0.2 mL (2 g)/mouse] was given only to the test group (18 mice). General behavior, clinical signs and feeding patterns were recorded. Blood and tissue samples were collected at intervals. Haematological parameters including platelet, red blood cell (RBC), white blood cell (WBC), packed cell volume (PCV), serum biochemistry including serum creatinine, serum glutamic-oxaloacetic transaminase (SGOT) and serum glutamic-pyruvic transaminase (SGPT) were determined. Organs for possible histopathological changes were examined. Results: Neither group exhibited alteration of behavior or reduction in food and water intake. Similarly, no significant changes in SGOT, SGPT and serum creatinine levels were detected in the test group. Histopathological organ changes were not observed in either group of mice except in three liver samples of the test group which had a mild focal necrosis. The platelet count (11.33±0.35)í105/µL (P=0.000 04) and the RBC count (7.97±0.61)í106/µL (P=0.000 03) were significantly increased in the test group compared to that of the controls. However, WBC count and PCV (%) values were not changed significantly in the test group. The platelet count in the test group started to increase significantly from Day 3 (3.4±0.18í105/µL), reaching almost a fourfold higher at Day 21 (11.3í105/µL), while it was 3.8í105/µL and 5.5í105/µL at Day 3 and Day 21 respectively in the control. Likewise, the RBC count in the test group increased from 6í106/µL to 9í106/ µL at Day 21 while it remained near constant in the control group (6í106/µL). Conclusions: Fresh C. papaya leaf extract significantly increased the platelet and RBC counts in the test group as compared to controls. Therefore, it is very

  4. Plecanatide and dolcanatide, novel guanylate cyclase-C agonists, ameliorate gastrointestinal inflammation in experimental models of murine colitis

    Kunwar; Shailubhai; Vaseem; Palejwala; Krishna; Priya; Arjunan; Sayali; Saykhedkar; Bradley; Nefsky; John; A; Foss; Stephen; Comiskey; Gary; S; Jacob; Scott; E; Plevy

    2015-01-01

    AIM: To evaluate the effect of orally administeredplecanatide or dolcanatide, analogs of uroguanylin, on amelioration of colitis in murine models.METHODS: The cyclic guanosine monophosphate(cG MP) stimulatory potency of plecanatide and dolcanatide was measured using a human colon carcinoma T84 cellbased assay. For animal studies all test agents were formulated in phosphate buffered saline. Sulfasalazine or 5-amino salicylic acid(5-ASA) served as positive controls. Effect of oral treatment with test agents on amelioration of acute colitis induced either by dextran sulfate sodium(DSS) in drinking water or by rectal instillation of trinitrobenzene sulfonic(TNBS) acid, was examined in BALB/c and/or BDF1 mice. Additionally, the effect of orally administered plecanatide on the spontaneous colitis in T-cell receptor alpha knockout(TCRα-/-) mice was also examined. Amelioration of colitis was assessed by monitoring severity of colitis, disease activity index and by histopathology. Frozen colon tissues were used to measure myeloperoxidase activity.RESULTS: Plecanatide and dolcanatide are structurally related analogs of uroguanylin, which is an endogenous ligand of guanylate cyclase-C(GC-C). As expected from the agonists of GC-C, both plecanatide and dolcanatide exhibited potent cG MP-stimulatory activity in T84 cells. Once-daily treatment by oral gavage with either of these analogs(0.05-0.5 mg/kg) ameliorated colitis in both DSS and TNBS-induced models of acute colitis, as assessed by body weight, reduction in colitis severity(P < 0.05) and disease activity index(P < 0.05). Amelioration of colitis by either of the drug candidates was comparable to that achieved by orally administered sulfasalazine or 5-ASA. Plecanatide also effectively ameliorated colitis in TCRα-/- mice, a model of spontaneous colitis. As dolcanatide exhibited higher resistance to proteolysis in simulated gastric and intestinal juices, it was selected for further studies. CONCLUSION: This is the first

  5. Folate-targeted paclitaxel-conjugated polymeric micelles inhibits pulmonary metastatic hepatoma in experimental murine H22 metastasis models

    Zhang Y

    2014-04-01

    Full Text Available Yan Zhang,1 Hui Zhang,2 Wenbin Wu,2 Fuhong Zhang,3,4 Shi Liu,3 Rui Wang,3 Yingchun Sun,1 Ti Tong,1 Xiabin Jing3 1Department of Thoracic Surgery, The Second Hospital of Jilin University, Changchun, Jilin, People's Republic of China; 2Department of Thoracic Surgery, Xuzhou Central Hospital, Xuzhou, Jiangsu, People's Republic of China; 3State Key Laboratory of Polymer Physics and Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun, Jilin, People's Republic of China; 4Department of Otolaryngology, The First Hospital of Lanzhou University, Lanzhou, Gansu, People's Republic of China Abstract: Hepatocellular carcinoma shows low response to most conventional chemotherapies; additionally, extrahepatic metastasis from hepatoma is considered refractory to conventional systemic chemotherapy. Target therapy is a promising strategy for advanced hepatoma; however, targeted accumulation and controlled release of therapeutic agents into the metastatic site is still a great challenge. Folic acid (FA and paclitaxel (PTX containing composite micelles (FA-M[PTX] were prepared by coassembling the FA polymer conjugate and PTX polymer conjugate. The main purpose of this study is to investigate the inhibitory efficacy of FA-M(PTX on the pulmonary metastasis of intravenously injected murine hepatoma 22 (H22 on BALB/c mice models. The lung metastatic burden of H22 were measured and tissues were analyzed by immunohistochemistry and histology (hematoxylin and eosin stain, followed by survival analysis. The results indicated that FA-M(PTX prevented pulmonary metastasis of H22, and the efficacy was stronger than pure PTX and simple PTX-conjugated micelles. In particular, the formation of lung metastasis colonies in mice was evidently inhibited, which was paralleled with the downregulated expression of matrix metalloproteinase-2 and matrix metalloproteinase-9. Furthermore, the mice bearing pulmonary metastatic hepatoma in the FA

  6. The Effect of Phosphatidylcholine and Deoxycholate Compound Injections to the Localized Adipose Tissue: An Experimental Study with a Murine Model

    Yongjoon Noh

    2012-09-01

    Full Text Available Background Phosphatidylcholine (PPC and deoxycholate (DCA compound has been recentlyused for the purpose of partial lipolysis and is valued for its efficacy and lower invasivenesscompared to liposuction and dermolipectomy used previously. In this article, the authors discussthe efficacy of the PPC dissolved in DCA via an experimental rat study model, along with suggestinga useful animal experimental model for the study of adipose tissue and lipolysis.Methods Bilateral inguinal fat pads of an experimental rat were elevated with the deep inferiorepigastric vessel as the sole vascular pedicle. Normal saline was injected on one side as acontrol group and a PPC and DCA compound was injected on the other side. After 4 days, therats were euthanized for microscopic tissue examination. The pathology was scored by a semiquantitativesystem in 4 categories: normal fat amount, fat necrosis, inflammatory activity,and stage of fibrosis. A Wilcoxon signed-rank test powered by SPSS packet program was usedfor statistical analysis and to determine significance.Results Microscopic examination was performed on the obtained samples, and theexperimental data of all four categories showed significant histologic differences compared tothe control group. All of the data also showed statistical significance by the Wilcoxon signedranktest (P<0.01.Conclusions In the inguinal fat pad rat model, the control group and the experimental grouphad a differed significantly in the amount of normal fat tissue, inflammation, necrosis, andfibrosis. We recommend the rat inguinal fat pad model used in this study, as it is likely to beuseful in related research.

  7. An Insight into the Behavior, Course and Kinetics of Acute Infection of Toxoplasma gondii Human RH Strain in Experimentally Infected Murine Model.

    Vikrant Sudan

    2014-03-01

    Full Text Available Toxoplasma gondii, an apicomplexan parasite, is capable of infecting a broad range of intermediate warm-blooded hosts including humans. The parasite seems to be capable of altering the natural behavior of the host to favor its transmission in the environment. The aim of this study was to evaluate the course, alterations in behavior along with normal kinetics of the abnormally induced experimental acute toxoplasmosis in murine models.Ten Swiss albino mice were intraperitoneally inoculated with 100 virulent RH strain tachyzoites and finally, the alterations in behavior were described and compared with other known alterations in humans and animals.The behavior and the other symptoms of the acute toxoplasmosis were recorded. Such mice showed typical symptoms like normal coat, severe ascites with pendulous abdomen and tachypnoea exhibited by resting fore legs either on walls of the cage, or nozzle of water bottle or other resting mice and yielded a creamy colored cloudy natured peritoneal fluid on aspiration.Finally the alterations in behavior were described and compared with other known alterations in humans and animals. The study has generated some important data related to possible causes of behavioral alterations and generation of suitable strategies for control of these alterations in behavior vis-à-vis better understanding of the effect of acute infection of parasite on normal behavior of infected intermediate host.

  8. Studies on the mechanisms responsible for inhibition of experimental metastasis of B16-F10 murine melanoma by pentoxifylline.

    Gude, R P; Binda, M M; Presas, H L; Klein-Szanto, A J; Bonfil, R D

    1999-01-01

    Pentoxifylline (PTX), a methylxanthine derivative widely used as a hemorheological agent in the treatment of peripheral vascular disease, was studied to unveil the mechanisms responsible for its inhibitory action on B16-F10 experimental metastasis. In vitro pretreatment of B16-F10 cells with noncytotoxic concentrations of PTX significantly inhibited their adhesion to reconstituted basement membrane Matrigel(R) and type IV collagen as well as the relative activity of secreted 92 kD metalloproteinase. However, PTX pretreatment of B16-F10 cells did not affect their in vitro invasiveness. Heterotypic organ adhesion assays carried out with B16-F10 cells and suspended organ tissues demonstrated that pretreatment with noncytotoxic concentrations of PTX of both, tumor cells or lung tissue, brought about a dose-dependent inhibition of melanoma cell adhesion to lung. Immunohistochemical studies using antibodies against CD31 adhesion molecule (PECAM-1) revealed that B16-F10 cells adhere to lung endothelial cells. Our results suggest that PTX may exert its inhibitory effect on tumor lodgment, and as a consequence of that on experimental metastases, through an inhibitory action on cell adhesion molecules. PMID:10087444

  9. Acute desipramine restores presynaptic cortical defects in murine experimental autoimmune encephalomyelitis by suppressing central CCL5 overproduction

    Di Prisco, Silvia; Merega, Elisa; Lanfranco, Massimiliano; Casazza, Simona; Uccelli, Antonio; Pittaluga, Anna

    2014-01-01

    Background and Purpose Altered glutamate exocytosis and cAMP production in cortical terminals of experimental autoimmune encephalomyelitis (EAE) mice occur at the early stage of disease (13 days post-immunization, d.p.i.). Neuronal defects were paralleled by overexpression of the central chemokine CCL5 (also known as RANTES), suggesting it has a role in presynaptic impairments. We propose that drugs able to restore CCL5 content to physiological levels could also restore presynaptic defects. Because of its efficacy in controlling CCL5 overexpression, desipramine (DMI) appeared to be a suitable candidate to test our hypothesis. Experimental Approach Control and EAE mice at 13 d.p.i. were acutely or chronically administered DMI and monitored for behaviour and clinical scores. Noradrenaline and glutamate release, cAMP, CCL5 and TNF-α production were quantified in cortical synaptosomes and homogenates. Peripheral cytokine production was also determined. Key Results Noradrenaline exocytosis and α2-adrenoeceptor-mediated activity were unmodified in EAE mice at 13 d.p.i. when compared with control. Acute, but not chronic, DMI reduced CCL5 levels in cortical homogenates of EAE mice at 13 d.p.i., but did not affect peripheral IL-17 and TNF-α contents or CCL5 plasma levels. Acute DMI caused a long-lasting restoration of glutamate exocytosis, restored endogenous cAMP production and impeded the shift from inhibition to facilitation of the CCL5-mediated control of glutamate exocytosis. Finally, DMI ameliorated anxiety-related behaviour but not motor activity or severity of clinical signs. Conclusions We propose DMI as an add-on therapy to normalize neuropsychiatric symptoms in multiple sclerosis patients at the early stage of the disease. PMID:24528439

  10. The efficacy of hydro alcoholic extract of Seidlitzia rosmarinus on experimental zoonotic cutaneous leishmaniasis lesions in murine model

    Maryam Ahmadi

    2014-11-01

    Full Text Available Objective: Leishmaniasis is one of the most important parasitic infectious diseases in the world. Since last century, many efforts have been made to control and treat the disease, but appropriate vaccines, pesticides and medicines are not available or even eligible. The purpose of this study was to evaluate the effect of hydro-alcoholic extract of Seidlitzia rosmarinus on the lesions of experimental Cutaneous Leishmaniasis (CL in Balb/c mice. Materials and Methods: The population study was 60 Ballb/c mice which divided to 6 groups, all infected with Leishmania major [MRHO/75/IR]. Soon after the ulcer started to appear in the early stage, a dose of provided herbal extract with 5, 10 and 15% concentration applied on each lesion. The surface area of the lesions measured during an interval of 10 days. Direct Giemsa stained smears prepared two and four weeks after treatment. Results: Increasing the mean size of the lesions was statistically significant compared to those in control group (p>0.001. Visceral Leishmaniasis (VL developed in all of the mice including the control group that received Eucerine alone. Survival rate in group receiving 15% S. rosmarinus extracts showed significantly higher  compared to mice in control group (p

  11. Cáncer experimental e inflamación sistémica en un modelo murino Systemic inflammation and experimental cancer in a murine model

    Juan Bruzzo

    2007-10-01

    both animals and human beings. In contrast, the relationship between cancer and systemic inflammation has been less studied. In this work, we demonstrated that the growth of the murine fibrosarcoma MC-C, was accompanied by manifestations of systemic inflammation, as demonstrated by an increase in both the number of circulating polymorphonuclear neutrophils (PMN and the serum concentration of the proinflammatory cytokines interleukin-1β (IL-1β, interleukin-6 (IL-6 and tumor necrosis factor-α (TNF-α and the acute phase proteins C reactive (CRP and serum A amyloid (SAA. Two temporally separate peaks of systemic inflammation were detected during tumor development. The first was displayed during the first week after tumor inoculation. The second peak began around day 14 and its intensity was proportional to tumor size. In mice bearing a large MC-C tumor, a high number of circulating PMN and myeloid precursors were evident. Most of these cells exhibited activation evidenced by an increased reactive oxygen species generation and high expression of the Gr1+/Mac1+ markers. Inoculation of thioglycolate -which generates a transient systemic inflammation- accelerated the growth of MC-C tumor and reciprocally, inhibition of such systemic inflammation by using indomethacin, prevented that enhancing effect. This suggests that the systemic inflammation that the tumor generates on its own, could be part of its growth strategy.

  12. Comparative Efficacies of Conventional Amphotericin B, Liposomal Amphotericin B (AmBisome), Caspofungin, Micafungin, and Voriconazole Alone and in Combination against Experimental Murine Central Nervous System Aspergillosis

    Clemons, Karl V.; Espiritu, Marife; Parmar, Rachana; Stevens, David A.

    2005-01-01

    Central nervous system (CNS) aspergillosis is a severe disease that responds poorly to current therapies. The current studies examined the efficacies of several antifungal agents alone or in combination with a murine model of CNS aspergillosis. Immunosuppressed mice were infected intracerebrally with Aspergillus fumigatus and treated with an amphotericin B preparation, an echinocandin, or voriconazole (VCZ) given alone or in combination. Monotherapy studies showed that micafungin (MICA), casp...

  13. A range finding protocol to support design for transcriptomics experimentation: examples of in-vitro and in-vivo murine UV exposure

    O. Bruning; W. Rodenburg; C.T. van Oostrom; M.J. Jonker; M. de Jong; R.J. Dekker; H. Rauwerda; W.A. Ensink; A. de Vries; T.M. Breit

    2014-01-01

    In transcriptomics research, design for experimentation by carefully considering biological, technological, practical and statistical aspects is very important, because the experimental design space is essentially limitless. Usually, the ranges of variable biological parameters of the design space a

  14. Biodistribution and pharmacokinetics of monoclonal antibody T1h and variant anti-CD6 murine 10D12 in healthy animals and in experimental arthritis model

    Biodistribution and pharmacokinetic of two radio labeled monoclonal antibodies was performed with the help of imaging techniques. Isotopic labeling was carried out by means of standardized methods. Pharmacokinetic evaluation was performed using the population approach and sparse data design. Introduction: Targeted therapy with monoclonal antibodies (MAb) is an efficient option for the treatment of rheumatoid arthritis. Th1 is a MAb anti human CD6 developed for the treatment of autoimmune disease and 10D12 is its counterpart anti murine CD6 developed as a pharmacological tool to get deep into the response mechanisms in animals models of rheumatoid arthritis.To investigate the behavior of both antibodies in the assay system, molecules were labeled with 125I to evaluate pharmacokinetic in healthy animals and with 99mTc to evaluate the antibody uptake in inflamed area of induced arthritis. Materials and methods: Antibodies were supplied by the Center of Molecular immunology. Iodination was performed by the iodogen method and technetium labeling was carried out directly by Schwarz method. Female C57BL6 from CENPALAB were used for experiments. Biodistribution and pharmacokinetic was performed by a sparse data design using the population approach. Uptake in region of inflammation was quantified by gammagraphy at the same time points of blood sampling. A compartmental model was build to quantify uptake kinetic. Pharmacokinetic profiles were analyzed using MONOLIX software version 4.2. Results: Minor pharmacokinetic differences were found between monoclonal antibodies labeled with 125I and 99mTc. As a humanized antibody, T1h shows a faster clearance than 10D12 and a biodistribution pattern reflecting preference for excretion mechanisms. The arthritis accumulation was not consistent with a targeted mediated uptake. On the other hand, radio labeled 10D12 shows an accumulation profile in arthritis with two peaks of maximum concentration representing an initial transit to

  15. Murine CD4+CD25- cells activated in vitro with PMA/ionomycin and anti-CD3 acquire regulatory function and ameliorate experimental colitis in vivo

    Majowicz Anna

    2012-12-01

    Full Text Available Abstract Background Induced regulatory T (iTreg lymphocytes show promise for application in the treatment of allergic, autoimmune and inflammatory disorders. iTreg cells demonstrate advantages over natural Treg (nTreg cells in terms of increased number of starting population and greater potential to proliferate. Different activation methods to generate iTreg cells result in iTreg cells that are heterogeneous in phenotype and mechanisms of suppression. Therefore it is of interest to explore new techniques to generate iTreg cells and to determine their physiological relevance. Methods Using phorbol myristate acetate (PMA/ionomycin and anti-CD3 activation of CD4+CD25- cells we generated in vitro functional CD4+CD25+ iTreg (TregPMA cells. Functionality of the generated TregPMA cells was tested in vivo in a mouse model of inflammatory bowel disease (IBD - CD45RB transfer colitis model. Results TregPMA cells expressed regulatory markers and proved to ameliorate the disease phenotype in murine CD45RB transfer colitis model. The body weight loss and disease activity scores for TregPMA treated mice were reduced when compared to diseased control group. Histological assessment of colon sections confirmed amelioration of the disease phenotype. Additionally, cytokine analysis showed decreased levels of proinflammatory colonic and plasma IL-6, colonic IL-1 β and higher levels of colonic IL-17 when compared to diseased control group. Conclusions This study identifies a new method to generate in vitro iTreg cells (TregPMA cells which physiological efficacy has been demonstrated in vivo.

  16. A Range Finding Protocol to Support Design for Transcriptomics Experimentation: Examples of In-Vitro and In-Vivo Murine UV Exposure

    Oskar Bruning; Wendy Rodenburg; van Oostrom, Conny T; Jonker, Martijs J; Mark de Jong; Dekker, Rob J.; Han Rauwerda; Ensink, Wim A.; Annemieke de Vries; Timo M Breit

    2014-01-01

    In transcriptomics research, design for experimentation by carefully considering biological, technological, practical and statistical aspects is very important, because the experimental design space is essentially limitless. Usually, the ranges of variable biological parameters of the design space are based on common practices and in turn on phenotypic endpoints. However, specific sub-cellular processes might only be partially reflected by phenotypic endpoints or outside the associated parame...

  17. A range finding protocol to support design for transcriptomics experimentation: examples of in-vitro and in-vivo murine UV exposure.

    Oskar Bruning

    Full Text Available In transcriptomics research, design for experimentation by carefully considering biological, technological, practical and statistical aspects is very important, because the experimental design space is essentially limitless. Usually, the ranges of variable biological parameters of the design space are based on common practices and in turn on phenotypic endpoints. However, specific sub-cellular processes might only be partially reflected by phenotypic endpoints or outside the associated parameter range. Here, we provide a generic protocol for range finding in design for transcriptomics experimentation based on small-scale gene-expression experiments to help in the search for the right location in the design space by analyzing the activity of already known genes of relevant molecular mechanisms. Two examples illustrate the applicability: in-vitro UV-C exposure of mouse embryonic fibroblasts and in-vivo UV-B exposure of mouse skin. Our pragmatic approach is based on: framing a specific biological question and associated gene-set, performing a wide-ranged experiment without replication, eliminating potentially non-relevant genes, and determining the experimental 'sweet spot' by gene-set enrichment plus dose-response correlation analysis. Examination of many cellular processes that are related to UV response, such as DNA repair and cell-cycle arrest, revealed that basically each cellular (sub- process is active at its own specific spot(s in the experimental design space. Hence, the use of range finding, based on an affordable protocol like this, enables researchers to conveniently identify the 'sweet spot' for their cellular process of interest in an experimental design space and might have far-reaching implications for experimental standardization.

  18. A range finding protocol to support design for transcriptomics experimentation: examples of in-vitro and in-vivo murine UV exposure.

    Bruning, Oskar; Rodenburg, Wendy; van Oostrom, Conny T; Jonker, Martijs J; de Jong, Mark; Dekker, Rob J; Rauwerda, Han; Ensink, Wim A; de Vries, Annemieke; Breit, Timo M

    2014-01-01

    In transcriptomics research, design for experimentation by carefully considering biological, technological, practical and statistical aspects is very important, because the experimental design space is essentially limitless. Usually, the ranges of variable biological parameters of the design space are based on common practices and in turn on phenotypic endpoints. However, specific sub-cellular processes might only be partially reflected by phenotypic endpoints or outside the associated parameter range. Here, we provide a generic protocol for range finding in design for transcriptomics experimentation based on small-scale gene-expression experiments to help in the search for the right location in the design space by analyzing the activity of already known genes of relevant molecular mechanisms. Two examples illustrate the applicability: in-vitro UV-C exposure of mouse embryonic fibroblasts and in-vivo UV-B exposure of mouse skin. Our pragmatic approach is based on: framing a specific biological question and associated gene-set, performing a wide-ranged experiment without replication, eliminating potentially non-relevant genes, and determining the experimental 'sweet spot' by gene-set enrichment plus dose-response correlation analysis. Examination of many cellular processes that are related to UV response, such as DNA repair and cell-cycle arrest, revealed that basically each cellular (sub-) process is active at its own specific spot(s) in the experimental design space. Hence, the use of range finding, based on an affordable protocol like this, enables researchers to conveniently identify the 'sweet spot' for their cellular process of interest in an experimental design space and might have far-reaching implications for experimental standardization. PMID:24823911

  19. Schistosoma mansoni Antigens Modulate Experimental Allergic Asthma in a Murine Model: a Major Role for CD4+ CD25+ Foxp3+ T Cells Independent of Interleukin-10▿

    Pacífico, Lucila G. G.; Marinho, Fábio A. V.; Fonseca, Cristina T; Michele M Barsante; Pinho, Vanessa; Sales-Junior, Policarpo A.; Luciana S Cardoso; Araújo, Maria Ilma; Carvalho, Edgar M; Cassali, Geovanni D; Teixeira, Mauro M; Oliveira, Sergio C.

    2008-01-01

    In areas where schistosomiasis is endemic, a negative correlation is observed between atopy and helminth infection, associated with a low prevalence of asthma. We investigated whether Schistosoma mansoni infection or injection of parasite eggs can modulate airway allergic inflammation in mice, examining the mechanisms of such regulation. We infected BALB/c mice with 30 S. mansoni cercariae or intraperitoneally injected 2,500 schistosome eggs, and experimental asthma was induced by ovalbumin (...

  20. In vivo evidence for CD4+ and CD8+ suppressor T cells in vaccination-induced suppression of murine experimental autoimmune thyroiditis

    In several experimental autoimmune diseases, including experimental autoimmune thyroiditis (EAT), vaccination with attenuated autoantigen-specific T cells has provided protection against subsequent induction of disease. However, the mechanism(s) of vaccination-induced suppression remains to be clarified. Since the authors have previously shown that suppression generated by pretreatment with mouse thyroglobulin (MTg) or thyroid-stimulating hormone in EAT is mediated by CD4+, not CD8+, suppressor T cells, they examined the role of T cell subsets in vaccination-induced suppression of EAT. Mice were vaccinated with irradiated, MTg-primed, and MTg-activated spleen cells and then challenged. Pretreatment with these cells suppressed EAT induced by immunization with MTg and adjuvant, but not by adoptive transfer of thyroiditogenic cells, suggesting a mechanism of afferent suppression. The activation of suppressor mechanisms did not require CD8+ cells, since mice depleted of CD8+ cells before vaccination showed reduced EAT comparable to control vaccinated mice. Furthermore, depletion of either the CD4+ or the CD8+ subset after vaccination did not significantly abrogate suppression. However, suppression was eliminated by the depletion of both CD4+ and CD8+ cells in vaccinated mice. These results provide evidence for the cooperative effects of CD4+ and CD8+ T cells in vaccination-induced suppression of EAT

  1. Suppression of NF-κB signaling and NLRP3 inflammasome activation in macrophages is responsible for the amelioration of experimental murine colitis by the natural compound fraxinellone

    Inflammatory bowel disease (IBD) affects millions of people worldwide. Although the etiology of this disease is uncertain, accumulating evidence indicates a key role for the activated mucosal immune system. In the present study, we examined the effects of the natural compound fraxinellone on dextran sulfate sodium (DSS)-induced colitis in mice, an animal model that mimics IBD. Treatment with fraxinellone significantly reduced weight loss and diarrhea in mice and alleviated the macroscopic and microscopic signs of the disease. In addition, the activities of myeloperoxidase and alkaline phosphatase were markedly suppressed, while the levels of glutathione were increased in colitis tissues following fraxinellone treatment. This compound also decreased the colonic levels of interleukin (IL)-1β, IL-6, IL-18 and tumor necrosis factor (TNF)-α in a concentration-dependent manner. These effects of fraxinellone in mice with experimental colitis were attributed to its inhibition of CD11b+ macrophage infiltration. The mRNA levels of macrophage-related molecules in the colon, including intercellular adhesion molecule 1 (ICAM1), vascular cell adhesion molecule 1 (VCAM1), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX2), were also markedly inhibited following fraxinellone treatment. The results from in vitro assays showed that fraxinellone significantly reduced lipopolysaccharide (LPS)-induced production of nitric oxide (NO), IL-1β and IL-18 as well as the activity of iNOS in both THP-1 cells and mouse primary peritoneal macrophages. The mechanisms responsible for these effects were attributed to the inhibitory role of fraxinellone in NF-κB signaling and NLRP3 inflammasome activation. Overall, our results support fraxinellone as a novel drug candidate in the treatment of colonic inflammation. - Highlights: • Fraxinellone, a lactone compound, alleviated DSS induced colitis. • The effects of fraxinellone were attributed to its inhibition on infiltrated

  2. Suppression of NF-κB signaling and NLRP3 inflammasome activation in macrophages is responsible for the amelioration of experimental murine colitis by the natural compound fraxinellone

    Wu, Xue-Feng; Ouyang, Zi-Jun; Feng, Li-Li; Chen, Gong; Guo, Wen-Jie; Shen, Yan; Wu, Xu-Dong; Sun, Yang, E-mail: yangsun@nju.edu.cn; Xu, Qiang, E-mail: molpharm@163.com

    2014-11-15

    Inflammatory bowel disease (IBD) affects millions of people worldwide. Although the etiology of this disease is uncertain, accumulating evidence indicates a key role for the activated mucosal immune system. In the present study, we examined the effects of the natural compound fraxinellone on dextran sulfate sodium (DSS)-induced colitis in mice, an animal model that mimics IBD. Treatment with fraxinellone significantly reduced weight loss and diarrhea in mice and alleviated the macroscopic and microscopic signs of the disease. In addition, the activities of myeloperoxidase and alkaline phosphatase were markedly suppressed, while the levels of glutathione were increased in colitis tissues following fraxinellone treatment. This compound also decreased the colonic levels of interleukin (IL)-1β, IL-6, IL-18 and tumor necrosis factor (TNF)-α in a concentration-dependent manner. These effects of fraxinellone in mice with experimental colitis were attributed to its inhibition of CD11b{sup +} macrophage infiltration. The mRNA levels of macrophage-related molecules in the colon, including intercellular adhesion molecule 1 (ICAM1), vascular cell adhesion molecule 1 (VCAM1), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX2), were also markedly inhibited following fraxinellone treatment. The results from in vitro assays showed that fraxinellone significantly reduced lipopolysaccharide (LPS)-induced production of nitric oxide (NO), IL-1β and IL-18 as well as the activity of iNOS in both THP-1 cells and mouse primary peritoneal macrophages. The mechanisms responsible for these effects were attributed to the inhibitory role of fraxinellone in NF-κB signaling and NLRP3 inflammasome activation. Overall, our results support fraxinellone as a novel drug candidate in the treatment of colonic inflammation. - Highlights: • Fraxinellone, a lactone compound, alleviated DSS induced colitis. • The effects of fraxinellone were attributed to its inhibition on

  3. Role of Omega-3 Polyunsaturated Fatty Acids in the Production of Prostaglandin E2 and Nitric Oxide during Experimental Murine Paracoccidioidomycosis

    S. C. Sargi

    2013-01-01

    Full Text Available There has recently been increased interest in the potential health effects of omega-3 polyunsaturated fatty acids on the immune system. Paracoccidioidomycosis is the most important endemic mycosis in Latin America. Macrophages have a fundamental role and act as first line of organism defense. The purpose of this study was to analyze the effect of n-3 fatty acids on the production of PGE2 and NO by mice infected with Pb18 and fed a diet enriched with LNA for 8 weeks. To study the effect of omega-3 fatty acids on macrophage activity during experimental paracoccidioidomycosis, mice were infected with Pb18 and fed a diet supplemented with LNA. PGE2 in the serum of animals was analyzed and NO in the supernatants of macrophages cultured and challenged in vitro with Pb18 was measured. Omega-3 fatty acids seemed to decrease the production of PGE2 in vivo in the infected group fed an LNA-supplemented diet during the 4th and 8th weeks of the experiment. At the same time, we observed an increase in synthesis of NO by peritoneal macrophages in this group. Omega-3 fatty acids thus appear to have an immunomodulatory effect in paracoccidioidomycosis.

  4. Expression of interleukin-17 in experimental models of murine mammary carcinoma%乳腺癌小鼠动物模型中白细胞介素-17的表达及其意义

    魏文; 李娟娟; 孙圣荣; 王卫星

    2011-01-01

    Objective To explore the impact of interleukin (IL) -17 expression on tumor growth in experimental models of murine mammary carcinoma and potential mechanisms. Methods Two murine cell lines, MA782/5S28102 and 4T1 were used to establish experimental models of murine mammary carcinoma. The IL-17 expression in tumor tissues derived from MA782-bearing mice or 4T1-bearing mice was detected in early and late stages of the tumor by Western blotting. The tumor cells and tumor-infiltrated-lymphocytes were separated from tumor tissues and cultured for 5 days with stimulation of PMA, anti-CD3 antibody and anti-CD28 antibody. The supernatants of culture media of stimulated tumor cells or tumor-infiltrated-lymphocytes were harvested and tested for IL-17 concentration by enzyme linked immunosorbent assay (ELISA). To evaluate the effect of IL-17 on the proliferation of tumor cells, 4T1 cells were culture in media with or without IL-17 and the cell number was counted on the day 5. For ire vivo assay, 4T1-bearing mice were injected with IL-17 or culture media via tail vein, and the tumor volume was measured. To assay the angiogenesis, the tumor tissues from 4T1-bearing mice with or without injection of IL-17 were stained with anti-CD31 antibody by immunohistochemistry. Results The IL-17 expression was significantly higher in late stage than in early stage of tumor in two experimental models. The tumor expression of IL-17 was secreted by tumor infiltrated lymphocytes (P <0.01). IL-17 could not increase the generation of tumor cells in vitro (1. 11 ±0. 11, 1. 28 ±0. 21 ,P >0. 05). But IL-17, injected into 4T1 -bearing mice, markedly enhanced in vivo tumor growth and significantly increased tumor vascularity (35. 79 ±9. 49, 13. 52 ±3. 55,P <0.01). Conclusion IL-17 in tumor tissue probably promotes tumor growth through enhancing angiogenesis.%目的 探讨小鼠乳腺癌动物模型中白细胞介素-17(IL-17)的表达及意义.方法 以MA782/5S28102及4T1细胞株建立两

  5. Local immunotherapy in experimental murine lung inflammation

    sprotocols

    2015-01-01

    Authors: Caroline Uebel, Sonja Koch, Anja Maier, Nina Sopel, Anna Graser, Stephanie Mousset & Susetta Finotto ### Abstract Innovative local immunotherapy for severe lung diseases such as asthma, chronic obstructive pulmonary disease or lung cancer requires a successful delivery to access the desired cellular target in the lung. An important route is the direct instillation into the airways in contrast to delivery through the digestive tract. This protocol details a method to deliv...

  6. Experimental study on IL-6,IL-10 and IL-23 expression in murine viral myocarditis%白细胞介素在小鼠病毒性心肌炎中表达的实验研究

    洪长江; 张园; 阮云军; 林朴卿

    2015-01-01

    目的:研究白细胞介素‐6(IL‐6)、IL‐10和IL‐23在小鼠病毒性心肌炎(VMC)中的表达,为临床治疗VMC提供相应的理论依据。方法将65只4周龄的雄性小鼠随机分成对照组25只、V M C组40只,分别设置0、1、2、4、6周5个时间点,其中VMC每亚组为8只,而对照组的亚组为5只,对VMC组各小鼠在腹腔内注射0.1 ml的病毒液,而对照组则注射0.1 ml的磷酸盐缓冲液(即PBS ),对比各时期两组小鼠心肌组织内IL‐6、IL‐10、IL‐23 mRNA水平及蛋白水平。结果实验组小鼠在感染柯萨奇病毒(CVB)后的2~4周内心肌组织产生的炎症反应最为明显,且 IL‐6、IL‐10及 IL‐23 mRNA 在各时期的表达量均显著高于对照组,差异均有统计学意义( P<0.05);实验组在0~6周各时期中的IL‐6、IL‐10及IL‐23蛋白水平均显著高于对照组相应时期,差异均有统计学意义(P<0.05)。结论 IL‐6、IL‐10及IL‐23可能与VMC发病过程有关,临床上应加以重视。%OBJECTIVE To investigate IL‐6 ,IL‐10 and IL‐23 expression in murine viral myocarditis (VMC) ,so as to provide theory information for clinical treatment of VMC .METHODS Totally 65 male four‐week‐old mice were randomly divided into the control group (25 cases) ,and VMC group (40 cases) ,in each subgroup of VMC had 8 cases ,while each subgroup of controls had 5 cases .Then five time points (0 week ,one week ,two weeks ,four weeks ,six weeks) were set .Each mouse in the VMC group was injected intraperitoneally with 0 .1 ml of the virus solution ,while the control group injected with 0 .1 ml phosphate buffered solution (PBS) .The mRNA and protein levels of IL‐6 ,IL‐10 and IL‐23 at each time point in the two groups were compared .RESULTS At 2 to 4 weeks after infection of coxsackie virus (CVB) in the experimental group ,the inflammatory reaction in myocardial tissues was the most obvious ,and

  7. Lactobacillus rhamnosus GG as an Effective Probiotic for Murine Giardiasis

    Nisha Goyal; Ram Prakash Tiwari; Geeta Shukla

    2011-01-01

    The gut microflora is an important constituent in the intestinal mucosal barrier and has been introduced as the concept of probiotic therapy that beneficially affects the host by improving its intestinal microbial balance. Therefore, the main objective of the study was to explore the protective potential of various lactobacilli strains for murine giardiasis. By experimentation, it was found that the probiotic supplementation of either Lactobacillus casei, L. acidophilus, L. plantarum, or L. r...

  8. Characterization of murine macrophages from bone marrow, spleen and peritoneum

    Wang Changqi; Yu Xiao; Cao Qi; Wang Ya; Zheng Guoping; Tan Thian Kui; Zhao Hong; Zhao Ye; Wang Yiping; Harris David CH

    2013-01-01

    Abstract Background Macrophages have heterogeneous phenotypes and complex functions within both innate and adaptive immune responses. To date, most experimental studies have been performed on macrophages derived from bone marrow, spleen and peritoneum. However, differences among macrophages from these particular sources remain unclear. In this study, the features of murine macrophages from bone marrow, spleen and peritoneum were compared. Results We found that peritoneal macrophages (PMs) app...

  9. Análisis de la resistencia inmune en un modelo murino experimental alimentado con dietas lipídicas e infectado con Listeria monocytogenes Analysis of the immune resistance in an experimental murine model fed dietary lipids and infected with Listeria monocytogenes

    M.ª A. Puertollano

    2004-11-01

    infectious diseases. The purpose of the present study was to determinate the immune status of mice fed dietary lipids and experimentally infected with a virulent strain of Listeria monocytogenes. Balb/c mice were divided into four groups and were fed with their respective diet: low fat diet (LF, 20%, olive oil diet (OO, 20%, fish oil diet (FO, 20% and hydrogenated coconut oil (HCO, 20%. Mice were fed for four weeks and infected with L. monocytogenes by endovenous route. Results have shown a survival reduction in mice fed a diet containing FO, as well as a significant increase in the number of viable bacteria from spleen. In addition, we have observed an increase in the bactericidal activity in peritoneal cells from OO group, although the invasion of L. monocytogenes in cells from this group was larger. Finally, a significant reduction of lymphocyte proliferation was observed in the group fed an FO diet, whereas natural killer (NK cell activity was not modified. These results indicate that dietary lipids constituted by polyunsaturated n-3 fatty acids reduce the murine immune resistance, whereas a diet constituted by OO-does not exert an immunosuppressor effect as relevant as FO diet, and it does not reduce the immune resistance leading to an efficient L. monocytogenes elimination.

  10. Identification of murine complement receptor type 2.

    Fingeroth, J D; Benedict, M A; Levy, D.N.; Strominger, J L

    1989-01-01

    A rabbit antiserum reactive with the human complement component C3d/Epstein-Barr virus receptor (complement receptor type 2, CR2) immunoprecipitates a Mr 155,000 murine B-cell surface antigen. The apparent molecular weight and cellular distribution of this murine antigen are similar to those of human CR2. Cells expressing the murine protein bind sheep erythrocytes coated with antibody and murine C1-C3d but do not bind Epstein-Barr virus at all. The monospecific antiserum to human CR2 together...

  11. Patogenesis of pipe-stem fibrosis of the liver (experimental observation on murine Schistosomiasis Patogenia da fibrose "pipe-stem" do fígado (observações experimentais na esquistossomose murina

    Zilton A. Andrade

    1987-09-01

    Full Text Available Mice infected with 30 cercariae of Schistosoma mansoni developed portal and septal fibrosis due to the massive and concentrated deposition of eggs in the periportal areas which occurred following the 16th week after infection. The lesion resembled pipe-stem fibrosis seen in human hepatosplenic schistosomiasis in the following characters: portal fibrosis interconnecting portal spaces as well as portal spaces and central canals; portal inflammation; periovular granulomas; vascular obstruction and telangiectasia. The liver parenchyma maintained its normal architecture. Vascular injection techniques with Indian ink and vinylite revealed that the portal system developed numerous dilated collateral venules coming from the large and medium-sized portal branches, about 10 weeks after schistosome infection. The lodging of schistosome eggs into these collaterals resulted in granulomatous inflammation and fibrosis along all the portal tracts, thus forming the pipe-stem lesion. Although not readily demonstrable grossly, the pipe-stem fibrosis of murine schistosomiasis has many similarities with the human lesion and can be considered to have the same basic pathogenesis.Camundongos infectados com 30 cercárias do Schistosoma mansoni desenvolveram fibrose porta em virtude de um depósito progressivo e concentrado de ovos na região periportal, o que aconteceu a partir da 16ª semana da infecção. Esta fibrose certas características da chamada fibrose "pipe-stem" do homem vista na forma hepatoesplênica da esquistossomose, tais como obstrução das radiculas porta, telangiectasia, conexão fibrosa entre espaços porta e entre estes e veias centrais, além de certo grau de fibrose septal, presença dos granulomas em várias fases evolutivas e reação inflamatória crônica difusa, enquanto o parênquima hepático mantinha a sua estrutura lobular normal. As técnicas de injeção vascular com tinta da China e com vinilite feitas no sistema porta permitiram a

  12. Direct demonstration of the infiltration of murine central nervous system by Pgp-1/CD44high CD45RB(low) CD4+ T cells that induce experimental allergic encephalomyelitis

    Zeine, R; Owens, T

    1992-01-01

    In experimental allergic encephalomyelitis (EAE), autoimmune T cells infiltrate the central nervous system (CNS) and initiate demyelinating pathology. We have used flow cytometry to directly analyse the migration to the CNS of MBP-reactive CD4+ T cells labelled with a lipophilic fluorescent dye...

  13. Murine Typhus and Febrile Illness, Nepal

    Zimmerman, Mark D.; Murdoch, David R.; Rozmajzl, Patrick J.; Basnyat, Buddha; Woods, Christopher W.; Richards, Allen L.; Belbase, Ram Hari; Hammer, David A.; Anderson, Trevor P.; Reller, L. Barth

    2008-01-01

    Murine typhus was diagnosed by PCR in 50 (7%) of 756 adults with febrile illness seeking treatment at Patan Hospital in Kathmandu, Nepal. Of patients with murine typhus, 64% were women, 86% were residents of Kathmandu, and 90% were unwell during the winter. No characteristics clearly distinguished typhus patients from those with blood culture–positive enteric fever.

  14. Tumor necrosis factor-mediated inhibition of interleukin-18 in the brain: a clinical and experimental study in head-injured patients and in a murine model of closed head injury.

    Shohami Esther

    2004-07-01

    Full Text Available Abstract Tumor necrosis factor (TNF and interleukin-(IL-18 are important mediators of neuroinflammation after closed head injury (CHI. Both mediators have been previously found to be significantly elevated in the intracranial compartment after brain injury, both in patients as well as in experimental model systems. However, the interrelation and regulation of these crucial cytokines within the injured brain has not yet been investigated. The present study was designed to assess a potential regulation of intracranial IL-18 levels by TNF based on a clinical study in head-injured patients and an experimental model in mice. In the first part, we investigated the interrelationship between the daily TNF and IL-18 cerebrospinal fluid levels in 10 patients with severe CHI for up to 14 days after trauma. In the second part of the study, the potential TNF-dependent regulation of intracerebral IL-18 levels was further characterized in an experimental set-up in mice: (1 in a standardized model of CHI in TNF/lymphotoxin-α gene-deficient mice and wild-type (WT littermates, and (2 by intracerebro-ventricular injection of mouse recombinant TNF in WT C57BL/6 mice. The results demonstrate an inverse correlation of intrathecal TNF and IL-18 levels in head-injured patients and a TNF-dependent inhibition of IL-18 after intracerebral injection in mice. These findings imply a potential new anti-inflammatory mechanism of TNF by attenuation of IL-18, thus confirming the proposed "dual" function of this cytokine in the pathophysiology of traumatic brain injury.

  15. Isolation of Murine Embryonic Hemogenic Endothelial Cells.

    Fang, Jennifer S; Gritz, Emily C; Marcelo, Kathrina L; Hirschi, Karen K

    2016-01-01

    The specification of hemogenic endothelial cells from embryonic vascular endothelium occurs during brief developmental periods within distinct tissues, and is necessary for the emergence of definitive HSPC from the murine extra embryonic yolk sac, placenta, umbilical vessels, and the embryonic aorta-gonad-mesonephros (AGM) region. The transient nature and small size of this cell population renders its reproducible isolation for careful quantification and experimental applications technically difficult. We have established a fluorescence-activated cell sorting (FACS)-based protocol for simultaneous isolation of hemogenic endothelial cells and HSPC during their peak generation times in the yolk sac and AGM. We demonstrate methods for dissection of yolk sac and AGM tissues from mouse embryos, and we present optimized tissue digestion and antibody conjugation conditions for maximal cell survival prior to identification and retrieval via FACS. Representative FACS analysis plots are shown that identify the hemogenic endothelial cell and HSPC phenotypes, and describe a methylcellulose-based assay for evaluating their blood forming potential on a clonal level. PMID:27341393

  16. Protective role of murine norovirus against Pseudomonas aeruginosa acute pneumonia.

    Thépaut, Marion; Grandjean, Teddy; Hober, Didier; Lobert, Pierre-Emmanuel; Bortolotti, Perrine; Faure, Karine; Dessein, Rodrigue; Kipnis, Eric; Guery, Benoit

    2015-01-01

    The murine norovirus (MNV) is a recently discovered mouse pathogen, representing the most common contaminant in laboratory mouse colonies. Nevertheless, the effects of MNV infection on biomedical research are still unclear. We tested the hypothesis that MNV infection could alter immune response in mice with acute lung infection. Here we report that co-infection with MNV increases survival of mice with Pseudomonas aeruginosa acute lung injury and decreases in vivo production of pro-inflammatory cytokines. Our results suggest that MNV infection can deeply modify the parameters studied in conventional models of infection and lead to false conclusions in experimental models. PMID:26338794

  17. Coxsackievirus B4 Infection of Murine Foetal Thymus Organ Cultures

    Brilot, F; Jaidane, H.; Geenen, Vincent; Hober, D

    2008-01-01

    The infection of foetal thymus with coxsackievirus B4 (CV-B4) E2 has been studied ex vivo by using CD-1 mice on foetal day 14, as a ready source of organs for experimentation to investigate the hypothesis of the role of thymic viral infections in the pathogenesis of type 1 diabetes. The replication of CV-B4 E2 in murine foetal thymus organ cultures has been demonstrated by evaluating the levels of positive- and negative-stranded viral RNA in cells by using a real-time quantitative RT-PCR meth...

  18. Multi-gene targeted antiangiogenic therapies for experimental corneal neovascularization

    Chen, Peng; Yin, Hongmei; Wang, Yao; Mi, Jing; He, Wenxiao; Xie, Lixin; Wang, Yiqiang

    2010-01-01

    Purpose To determine the effectiveness of multigene-based anti-angiogenic gene therapies for experimental murine corneal neovascularization (corneal NV). Methods Recombinant retroviral vectors encoding murine endostatin (mEndo), murine-soluble vascular endothelial growth factor receptor-2 (msFlk-1), or murine-soluble Tie2 (msTie2) were constructed and packaged in PT67 cells. Viral titers were determined by infection of NIH3T3 cells. Expressions of mEndo, msFlk-1, and msTie2 were confirmed by ...

  19. Chemoimmunotherapy of murine bladder cancer.

    Stogdill, B J; Lamm, D L; Livingston, R B

    1981-11-01

    The lethality of invasive transitional cell carcinoma (TCC) has prompted a search for effective, minimally toxic, adjuvant therapy. Such agents were evaluated in a murine bladder cancer (MBT2) model which parallels the clinical disease. One hundred C3H/He mice were inoculated i.d. with 2.5 x 10(4) viable MBT2 tumor cells and randomized to receive either normal saline (control), cis-Platinum (CPT), cyclophosphamide (CY), methotrexate (MTX), BCG, (CY + MTX), or (CY + MTX + BCG). Chemotherapy was given intraperitoneally weekly starting on day 7 after inoculation. Immunotherapy was given intralesionally on days 1 and 10 only. All mice were treated for 5 weeks followed by 5 weeks of observation. At 5 weeks, tumors of mice receiving cyclophosphamide alone or either of the combinations of therapy were smaller (P less than 0.01) than tumors of controls or other single agents alone. Each regimen increased survival, but only the combination regimen increase survival significantly (P less than 0.01). In the doses and schedule used in this model. Combination chemotherapy and chemoimmunotherapy significantly delay tumor growth and increase duration of survival (P less than 0.01) when compared with controls or single agent groups. PMID:7298287

  20. Apoptosis in irradiated murine tumors.

    Stephens, L C; Ang, K K; Schultheiss, T E; Milas, L; Meyn, R E

    1991-09-01

    Early radiation responses of transplantable murine ovarian (OCaI) and hepatocellular (HCaI) carcinomas were examined at 6, 24, 48, 96, and 144 h after single photon doses of 25, 35, or 45 Gy. Previous studies using tumor growth delay and tumor radiocurability assays had shown OCaI tumors to be relatively radiosensitive and HCaI tumors to be radioresistant. At 6 h, approximately 20% of nuclei in OCaI tumors showed aberrations characteristic of cell death by apoptosis. This contrasted to an incidence of 3% in HCaI tumors. Mitotic activity was eliminated in OCaI tumors but was only transiently suppressed in HCaI tumors. At 24-96 h, OCaI tumors continued to display apoptosis and progressive necrosis, whereas HCaI tumors responded by exhibiting marked pleomorphism. Factors other than mitotic activity may influence tumor radiosensitivity, and one of these may be susceptibility to induction of apoptosis (programmed cell death), because this was a prominent early radiation response by the radiosensitive OCaI tumors. PMID:1886987

  1. Histopathological characterization of a syngeneic orthotopic murine bladder cancer model

    Daher C. Chade

    2008-03-01

    Full Text Available PURPOSE: We developed and characterized by histopathology and immunohistochemistry a syngeneic murine bladder tumor model derived from the MB49 tumor cell line. MATERIALS AND METHODS: Bladder tumor implantation was achieved by intravesical instillation of 5 x 10(5 MB49 tumor cells in C57BL/6 mice. A chemical lesion of the bladder was performed in order to promote intravesical tumor implantation. The bladder wall lesion was accomplished by transurethral instillation of silver nitrate (AgNO3. After 15 days, the animals were sacrificed, examined macroscopically for intravesical tumor and bladder weight. Histology and immunohistochemistry were performed using cytokeratin 7 (CK7, carcinoembrionic antigen (Dako-CEA, p53 and c-erbB2 oncoprotein (Her2/neu. RESULTS: Twenty-nine out of 30 animals (96.7% developed intravesical tumors in a 15-day period. Macroscopically, the mean bladder weight was 0.196g (0.069-0.538g, 10 to 15 times the normal bladder weight. The immunohistochemical analysis showed significant membrane expression of CEA and CK7: a similar finding for human urothelial cancer. We also characterized absence of expression of p53 and anti-Her2/neu in the murine model. CONCLUSIONS: High tumor take rates were achieved by using the chemical induction of the bladder tumor. Although electric cauterization is widely described in the literature for syngeneic orthotopic animal models, the technique described in this study represents an alternative for intravesical bladder tumor implantation. Moreover, the histopathology and immunohistochemical analysis of the murine bladder tumor model derived from the MB49 cell line showed a resemblance to human infiltrating urothelial carcinoma, allowing clinical inference from experimental immunotherapy testing.

  2. Three-dimensional in vivo imaging of the murine liver: a micro-computed tomography-based anatomical study.

    Teresa Fiebig

    Full Text Available Various murine models are currently used to study acute and chronic pathological processes of the liver, and the efficacy of novel therapeutic regimens. The increasing availability of high-resolution small animal imaging modalities presents researchers with the opportunity to precisely identify and describe pathological processes of the liver. To meet the demands, the objective of this study was to provide a three-dimensional illustration of the macroscopic anatomical location of the murine liver lobes and hepatic vessels using small animal imaging modalities. We analysed micro-CT images of the murine liver by integrating additional information from the published literature to develop comprehensive illustrations of the macroscopic anatomical features of the murine liver and hepatic vasculature. As a result, we provide updated three-dimensional illustrations of the macroscopic anatomy of the murine liver and hepatic vessels using micro-CT. The information presented here provides researchers working in the field of experimental liver disease with a comprehensive, easily accessable overview of the macroscopic anatomy of the murine liver.

  3. Bone marrow stromal elements in murine leukemia

    A study of bone marrow stromal elements in murine acute myeloid leukemia (AML) was carried out. Our previous studies had indicated marrow stromal deficiency in murine AML. In the current investigation, separate stromal cells were cultured and the results obtained have shown that, while marrow stromal macrophages are normal in leukemia and express adequate amounts of IL-1, the fibroblasts are markedly reduced. However, if sufficient fibroblasts are pooled in vitro, they produce adequate amounts of CSF. Test of TNFα in leukemic cells CM, as possible cause of marrow stromal inhibition in leukemia, had not disclosed this cytokine. Further, it was observed that total body lethal irradiation of leukemic mice aggravates the stromal deficiency, confirming results of our previous investigations. It is concluded that bone marrow stromal deficiency in murine AML is due to decreased fibroblasts and, implicity, reduced CSF production. (author)

  4. Establishment of murine model of radiation-induced oral mucositis

    Objective: To establish a murine model of radiation-induced oral mucositis. Methods: Left-sided buccal mucosa of 70 Sprague-Dawley rats were irradiated with X-rays (10 Gy/d), then six rats, selected randomly, were sacrificed at the 2nd, 4th, 6th, 8th, 14th, and 21st day after the irradiation, left-sided buccal mucosa were excised, corresponding irradiation dose for the selected rats were 20, 40, 60 and 80 Gy following radiation. The right buccal mucosa was excised and treated as its auto-controls tissue. Results: Erythema was observed in the left-sided buecal mucosa in rates irradiated with X-rays of 60 Gy; a single or more ulcers observed in rates irradiated with 80 Gy X-rays; a large area of ulcer was observed at 4th day after 80 Gy irradiation. The radiation-induced ulcer in buccal mucosa was essentially recovered after about 2 weeks following 80 Gy irradiation. Conclusion: This murine model of radiation-induced oral mucositis was useful and practical in experimental studies. (authors)

  5. Murine Typhus in Child, Yucatan, Mexico

    Zavala-Castro, Jorge E.; Zavala-Velázquez, Jorge E.; Uicab, Justo Eduardo Sulú

    2009-01-01

    A case of murine typhus in Yucatan was diagnosed in a child with nonspecific signs and symptoms. The finding of Rickettsia typhi increases the number of Rickettsia species identified in Yucatan and shows that studies are needed to determine the prevalence and incidence of rickettsioses in Mexico.

  6. Reemergence of Murine Typhus in the US

    2015-04-21

    Dr. Lucas Blanton discusses the Reemergence of Murine Typhus in Galveston Texas in 2013.  Created: 4/21/2015 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 4/27/2015.

  7. Multiphoton Imaging of Ultrasound Bioeffects in the Murine Brain

    Raymond, Scott; Skoch, Jesse; Bacskai, Brian; Hynynen, Kullervo

    2006-05-01

    The purpose of this study was to demonstrate the feasibility of multiphoton imaging in the murine brain during exposure to ultrasound. Our experimental setup coupled ultrasound through the ventral surface of the mouse while allowing imaging through a cranial window from the dorsal surface. Field attenuation was estimated by scanning the field after insertion of a freshly sacrificed mouse; beam profile and peak position were preserved, suggesting adequate targeting for imaging experiments. C57 mice were imaged with a Biorad multiphoton microscope while being exposed to ultrasound (f = 1.029 MHz, peak pressure ˜ 200 kPa, average power ˜ 0.18 W) with IV injection of Optison. We observed strong vasoconstriction coincident with US and Optison, as well as permeabilization of the blood-brain barrier.

  8. Murine granulated metrial gland cells are susceptible to Chlamydia psittaci infection in vivo.

    Sánchez, J.; Buendía, A.J.; Salinas, J.; Bernabé, A.; Rodolakis, A; Stewart, I J

    1996-01-01

    Granulated metrial gland (GMG) cells are the most numerous lymphoid cells in the uteroplacental unit in rodent pregnancy. In an experimental murine model of abortion-causing infection, we have studied the responses of GMG cells to Chlamydia psittaci. Chlamydial inclusions have been found within GMG cells, both in apparently healthy cells and in cells with degenerative changes. Establishing the existence of GMG cells infected by C. psittaci opens a new and interesting chapter in the study of t...

  9. Development and characterization of Histoplasma capsulatum-reactive murine T-cell lines and clones.

    Deepe, G S; Smith, J G; Sonnenfeld, G; Denman, D.; Bullock, W E

    1986-01-01

    Experimental studies have suggested that antigen-specific T lymphocytes are important mediators of resistance to infection with the pathogenic fungus Histoplasma capsulation. To gain a better understanding of the role of T lymphocytes, we developed murine T-cell lines and clones that recognized Histoplasma antigens. These T cells were of the helper/inducer phenotype (Thy-1.2+ Lyt-1+ L3T4+ Lyt-2-) and exerted multiple immunological functions. T-cell lines and 12 clones proliferated vigorously ...

  10. Biomarkers of Disease and Treatment in Murine and Cynomolgus Models of Chronic Asthma

    Jennifer Louten,; Mattson, Jeanine D.; Maria-Christina Malinao; Ying Li; Claire Emson; Felix Vega; Robert L. Wardle; Scott, Michael R.; Fick, Robert B.; McClanahan, Terrill K.; Rene de Waal Malefyt; Maribel Beaumont

    2012-01-01

    Background Biomarkers facilitate early detection of disease and measurement of therapeutic efficacy, both at clinical and experimental levels. Recent advances in analytics and disease models allow comprehensive screening for biomarkers in complex diseases, such as asthma, that was previously not feasible. Objective Using murine and nonhuman primate (NHP) models of asthma, identify biomarkers associated with early and chronic stages of asthma and responses to steroid treatment. Methods The tot...

  11. Simultaneous administration of vitamin A and DTP vaccine modulates the immune response in a murine cerebral malaria model

    Hein-Kristensen, L; Jørgensen, M J; Ravn, H;

    2010-01-01

    -tetanus-pertussis (DTP) vaccine may increase mortality from non-targeted diseases. We investigated the non-targeted effect of pretreatment with VAS and DTP vaccine in a murine model of experimental cerebral malaria. Our a priori hypothesis was that VAS/DTP would aggravate the infection. We found that the effect of VAS...

  12. Dose-effect relationship of apoptosis induced by fission-neutron in murine thymocytes

    Objective: To investigate the effectiveness of high LET fission-neutron to induce apoptosis in murine thymocytes and to compare it with that of low LET 60Co γ-ray. Methods: Apoptosis induction was studied qualitatively by light and transmission electron microscopy and DNA gel electrophoresis,also quantitatively by flow cytometry(FCM) and diphenylamine (DPA)methods. Results: DNA ladders of murine thymocytes were detectable, the typical apoptosis of thymocytes could be observed morphologically by means of light and electron microscopy at 6 h after fission-neutron irradiation with doses ranging from 0.5 to 5.0 Gy, meanwhile the percentages of apoptosis increased with increasing doses. After exposure to γ-rays with doses ranging from 1.0 to 30 Gy, the experimental results were similar to those from neutron radiation. The incidence of apoptosis peaked at about 20 Gy, the percentages did not increase further when doses increased. Conclusion: Apoptosis of murine thymocytes can be induced when mice are exposed to either fission-neutron (0.5-5.0 Gy) or to γ-ray (1-30 Gy). Although the relationship between apoptosis and radiation doses is similar, the percentage of apoptosis induced by neutron irradiation is higher than that induced by γ-irradiation. The RBE values of fission-neutron for inducing apoptosis murine thymocytes are 2.09 (by FCM method) and 2.37 (by DPA method), respectively. These results also suggest that fission-neutron-induced murine immune tissue is more severe than that induced by γ-rays at several hours post-irradiation and this might be the basis for heavy damage to immune tissues induced by fission-neutron-irradiation in later period

  13. Retroviral Transduction of Murine Primary T Lymphocytes

    Lee, James; Sadelain, Michel; Brentjens, Renier

    2016-01-01

    Summary In comparison to human T cells, efficient retroviral gene transfer and subsequent expansion of murine primary T cells is more difficult to achieve. Herein, we describe an optimized gene transfer protocol utilizing an ecotropic viral vector to transduce primary murine T cells activated with magnetic beads coated with agonistic anti-CD3 and CD28 antibodies. Activated T cells are subsequently centrifuged (spinoculated) on RetroNectin-coated tissue culture plates in the context of retroviral supernatant. Variables found to be critical to high gene transfer and subsequent efficient T cell expansion included CD3/CD28 magnetic bead to cell ratio, time from T cell activation to initial spinoculation, frequency of T cell spinoculation, interleukin-2 concentration in the medium, and the initial purity of the T cell preparation. PMID:19110621

  14. Murine Typhus: Clinical and epidemiological aspects

    Gaspar Peniche Lara

    2012-06-01

    Full Text Available Rickettsia typhi is an intracellular bacteria who causes murine typhus. His importance is reflected in the high frequency founding specific antibodies against R. typhi in several worldwide seroepidemiological studies, the seroprevalence ranging between 3-36%. Natural reservoirs of Rickettsia typhi are rats (some species belonging the Rattus Genus and fleas (Xenopsylla cheopis are his vector. This infection is associated with overcrowding, pollution and poor hygiene. Typically presents fever, headache, rash on trunk and extremities, in some cases may occur organ-specific complications, affecting liver, kidney, lung or brain. Initially the disease is very similar to other diseases, is very common to confuse the murine typhus with Dengue fever, therefore, ignorance of the disease is a factor related to complications or non-specific treatments for the resolution of this infection. This paper presents the most relevant information to consider about the rickettsiosis caused by Rickettsia typhi.

  15. Eliminating Murine Norovirus by Cross-Fostering

    Buxbaum, Laurence U.; DeRitis, Pierina C.; Chu, Niansheng; Conti, Pierre A.

    2011-01-01

    Murine norovirus (MNV) is a newly discovered and extremely prevalent pathogen of laboratory mouse colonies. MNV causes severe disease in some immunocompromised mouse strains and can cause persistent infections even in immunocompetent mice. Despite the fact that immunocompetent mice are generally asymptomatic, the possibility that MNV infection might alter immune responses makes its eradication a potentially useful goal for many facilities. Initial attempts by others to use a strategy of testi...

  16. Signaling pathways regulating murine pancreatic development

    Serup, Palle

    2012-01-01

    The recent decades have seen a huge expansion in our knowledge about pancreatic development. Numerous lineage-restricted transcription factor genes have been identified and much has been learned about their function. Similarly, numerous signaling pathways important for pancreas development have...... been identified and the specific roles have been investigated by genetic and cell biological methods. The present review presents an overview of the principal signaling pathways involved in regulating murine pancreatic growth, morphogenesis, and cell differentiation....

  17. Cell-free translation of murine coronavirus RNA.

    Leibowitz, J L; Weiss, S.R.; Paavola, E; Bond, C W

    1982-01-01

    The coding assignments of the intracellular murine hepatitis virus-specific subgenomic RNA species and murine hepatitis virion RNA have been investigated by cell-free translation. The six murine hepatitis virus-specific subgenomic RNAs were partially purified by agarose gel electrophoresis and translated in an mRNA-dependent rabbit reticulocyte lysate, and the cell-free translation products were characterized by gel electrophoresis, immunoprecipitation, and tryptic peptide mapping. These stud...

  18. Murine fundus fluorescein angiography: An alternative approach using a handheld camera.

    Ehrenberg, Moshe; Ehrenberg, Scott; Schwob, Ouri; Benny, Ofra

    2016-07-01

    In today's modern pharmacologic approach to treating sight-threatening retinal vascular disorders, there is an increasing demand for a compact, mobile, lightweight and cost-effective fluorescein fundus camera to document the effects of antiangiogenic drugs on laser-induced choroidal neovascularization (CNV) in mice and other experimental animals. We have adapted the use of the Kowa Genesis Df Camera to perform Fundus Fluorescein Angiography (FFA) in mice. The 1 kg, 28 cm high camera has built-in barrier and exciter filters to allow digital FFA recording to a Compact Flash memory card. Furthermore, this handheld unit has a steady Indirect Lens Holder that firmly attaches to the main unit, that securely holds a 90 diopter lens in position, in order to facilitate appropriate focus and stability, for photographing the delicate central murine fundus. This easily portable fundus fluorescein camera can effectively record exceptional central retinal vascular detail in murine laser-induced CNV, while readily allowing the investigator to adjust the camera's position according to the variable head and eye movements that can randomly occur while the mouse is optimally anesthetized. This movable image recording device, with efficiencies of space, time, cost, energy and personnel, has enabled us to accurately document the alterations in the central choroidal and retinal vasculature following induction of CNV, implemented by argon-green laser photocoagulation and disruption of Bruch's Membrane, in the experimental murine model of exudative macular degeneration. PMID:27260483

  19. Murine erythrocytes contain high levels of lysophospholipase activity

    Kamp, J.A.F. op den; Roelofsen, B.; Sanderink, G.; Middelkoop, E.; Hamer, R.

    1984-01-01

    Murine erythrocytes were found to be unique in the high levels of lysophospholipase activity in the cytosol of these cells. The specific activity of the enzyme in the cytosol of the murine cells is 10-times higher than in the cytosol of rabbit erythrocytes and approximately three orders of magnitude

  20. Exploring the translational disconnect between the murine and human inflammatory response: analysis of LPS dose–response relationship in murine versus human cell lines and implications for translation into murine models of sepsis

    McCarron EP

    2015-10-01

    Full Text Available Eamon P McCarron,1 Dominic P Williams,1 Daniel J Antoine,1 Anja Kipar,2 Jana Lemm,3 Sebastian Stehr,3 Ingeborg D Welters,4 1Department of Clinical and Molecular Pharmacology, Centre for Drug Safety Science, Institute of Translational Medicine, 2Department of Veterinary Pathology, University of Liverpool, Liverpool, UK; 3Department of Anaesthesiology and Intensive Care Medicine, Jena University Hospital, Jena, Germany; 4Department of Obesity and Endocrinology, Institute of Ageing and Chronic Disease, University of Liverpool, Liverpool, UK Background: Inflammation forms an important part of the human innate immune system and is largely dependent on the activation of the "classical" NF-κB pathway through Toll-like receptors (TLRs. Understanding this has allowed researchers to explore roles of therapeutic targets in managing conditions such as sepsis. Recapitulating an inflammatory response using lipopolysaccharide (LPS, a "sterile" technique, can provide information that is dissimilar to the clinical condition. By examining NF-κB activation (through immunoblotting of the p65 subunit in two separate cell lines (murine and human and analyzing two murine models of sepsis (intraperitoneal [IP] LPS and IP stool inoculation, an evaluation of the translational disconnect between experimental and clinical sepsis can be made. Methods: THP-1 (human cells and RAW 264.7 (murine cells were dosed with concentrations of LPS (human, 1 pg/mL to 100 ng/mL; murine, 30 pg/mL to 1,000 ng/mL and nuclear actin and p65 were immunoblotted to measure changes in nuclear density. In vivo, C57BL/6 mice received either IP injection of stool suspension (5 µL/g or LPS (25 mg/kg or saline (1 mL/kg. Animals were culled at 6 hours and tissues were analyzed. Results: An increase in basal p65:actin density in THP-1 cells (mean 0.214, standard error of the mean 0.024 was seen at doses as small as 0.1 ng/mL (0.519±0.064. In contrast to RAW 264.7 cells, basal increases (0.170±0

  1. Splenectomy normalizes hematocrit in murine polycythemia vera.

    Jan-Rung Mo

    Full Text Available Splenic enlargement (splenomegaly develops in numerous disease states, although a specific pathogenic role for the spleen has rarely been described. In polycythemia vera (PV, an activating mutation in Janus kinase 2 (JAK2(V617 induces splenomegaly and an increase in hematocrit. Splenectomy is sparingly performed in patients with PV, however, due to surgical complications. Thus, the role of the spleen in the pathogenesis of human PV remains unknown. We specifically tested the role of the spleen in the pathogenesis of PV by performing either sham (SH or splenectomy (SPL surgeries in a murine model of JAK2(V617F-driven PV. Compared to SH-operated mice, which rapidly develop high hematocrits after JAK2(V617F transplantation, SPL mice completely fail to develop this phenotype. Disease burden (JAK2(V617 is equivalent in the bone marrow of SH and SPL mice, however, and both groups develop fibrosis and osteosclerosis. If SPL is performed after PV is established, hematocrit rapidly declines to normal even though myelofibrosis and osteosclerosis again develop independently in the bone marrow. In contrast, SPL only blunts hematocrit elevation in secondary, erythropoietin-induced polycythemia. We conclude that the spleen is required for an elevated hematocrit in murine, JAK2(V617F-driven PV, and propose that this phenotype of PV may require a specific interaction between mutant cells and the spleen.

  2. Murine Typhus: Clinical and epidemiological aspects

    Gaspar Peniche Lara

    2012-06-01

    Full Text Available 14.00 Normal 0 21 false false false ES-CO X-NONE X-NONE Rickettsia typhi is an intracellular bacteria who causes murine typhus. His importance is reflected in the high frequency founding specific antibodies against R. typhi in several worldwide seroepidemiological studies, the seroprevalence ranging between 3-36%. Natural reservoirs of Rickettsia typhi are rats (some species belonging the Rattus Genus and fleas (Xenopsylla cheopis are his vector. This infection is associated with overcrowding, pollution and poor hygiene. Typically presents fever, headache, rash on trunk and extremities, in some cases may occur organ-specific complications, affecting liver, kidney, lung or brain. Initially the disease is very similar to other diseases, is very common to confuse the murine typhus with Dengue fever, therefore, ignorance of the disease is a factor related to complications or non-specific treatments for the resolution of this infection. This paper presents the most relevant information to consider about the rickettsiosis caused by Rickettsia typhi.

  3. A Rapid Murine Coma and Behavior Scale for Quantitative Assessment of Murine Cerebral Malaria

    Carroll, Ryan W.; Mark S Wainwright; KIM, KWANG-YOUN; Kidambi, Trilokesh; Gómez, Noé D.; Taylor, Terrie; Haldar, Kasturi

    2010-01-01

    Background Cerebral malaria (CM) is a neurological syndrome that includes coma and seizures following malaria parasite infection. The pathophysiology is not fully understood and cannot be accounted for by infection alone: patients still succumb to CM, even if the underlying parasite infection has resolved. To that effect, there is no known adjuvant therapy for CM. Current murine CM (MCM) models do not allow for rapid clinical identification of affected animals following infection. An animal m...

  4. Heterohybridoma for the production of non murine monoclonal antibodies

    Kh.Victoria Chanu and M. Ayub Ali

    Full Text Available Hybridoma technology described by kohler and Milstein produce only mouse immunoglobulins. Such immunoglobulins have limited use due to its negative side effects such as the recipient’s immune response. The production of a non murine monoclonal antibody to combat the problems of murine monoclonal antibody is again difficult due to the lack of a suitable myeloma cell line. Heterohybridoma formed by the fusion of lymphocyte of one species with the myeloma cell of a different species is the solution, which can be used for the production of non murine monoclonal antibodies. [Veterinary World 2010; 3(8.000: 390-392

  5. Murine Typhus in Southern Taiwan during 1992–2009

    Chang, Ko; Chen, Yen-Hsu; Lee, Nan-Yao; Lee, Hsin-Chun; Lin, Chun-Yu; Tsai, Jih-Jin; Lu, Po-Liang; Chen, Tun-Chieh; Hsieh, Hsiao-Chen; Lin, Wei-Ru; Lai, Ping-Chang; Chang, Chia-Ming; Wu, Chi-Jung; Lai, Chung-Hsu; Ko, Wen-Chien

    2012-01-01

    Clinical information regarding murine typhus in Taiwan is limited. In this study, 81 cases of serologically documented murine typhus during 1992–2009 at four referral hospitals in southern Taiwan were analyzed. There was a significant correlation between average environmental temperature and case numbers of murine typhus (r = 0.747, P = 0.005). Acute hepatitis was found in 67% of cases, and hyperbilirubinemia (serum total bilirubin ≥ 23.9 μmol/L) was found in 38%. The intervals between the in...

  6. Proliferative capacity of murine hematopoietic stem cells

    The present study demonstrates a decrease in self-renewal capacity with serial transfer of murine hematopoietic stem cells. Production of differentiated cell progeny is maintained longer than stem cell self-renewal. In normal animals the capacity for self-renewal is not decreased with increasing donor age. The stem cell compartment in normal animals, both young and old, appears to be proliferatively quiescent. After apparent recovery from the alkylating agent busulfan, the probability of stem cell self-renewal is decreased, there is a permanent defect in the capacity of the bone marrow for serial transplantation, and the stem cells are proliferatively active. These findings support a model of the hematopoietic stem cell compartment as a continuum of cells with decreasing capacities for self-renewal, increasing likelihood for differentiation, and increasing proliferative activity. Cells progress in the continuum in one direction and such progression is not reversible

  7. Induction of Th1Immune responses following laser ablation in a murine model of colorectal liver metastases

    Muralidharan Vijayaragavan; Nikfarjam Mehrdad; Malcontenti-Wilson Caterina; Fifis Theodora; Lin Wen; Nguyen Linh; Christophi Christopher

    2011-01-01

    Abstract Background Preliminary experimental studies have suggested that the in situ destruction of tumor tissue by local laser ablation (LA) may also stimulate host immunity against cancer. We investigated local and systemic induction of immune responses after laser ablation in the setting of residual tumor. Methods A murine colorectal cancer (CRC) liver metastasis model was used. Selected tumors of liver CRC bearing mice and livers of mice without tumor induction were treated with LA. Liver...

  8. Exploring the translational disconnect between the murine and human inflammatory response: analysis of LPS dose–response relationship in murine versus human cell lines and implications for translation into murine models of sepsis

    McCarron, Eamon P; Williams, Dominic P; Antoine, Daniel J; Kipar, Anja; Lemm, Jana; Stehr, Sebastian; Welters, Ingeborg D

    2015-01-01

    Background Inflammation forms an important part of the human innate immune system and is largely dependent on the activation of the “classical” NF-κB pathway through Toll-like receptors (TLRs). Understanding this has allowed researchers to explore roles of therapeutic targets in managing conditions such as sepsis. Recapitulating an inflammatory response using lipopolysaccharide (LPS), a “sterile” technique, can provide information that is dissimilar to the clinical condition. By examining NF-κB activation (through immunoblotting of the p65 subunit) in two separate cell lines (murine and human) and analyzing two murine models of sepsis (intraperitoneal [IP] LPS and IP stool inoculation), an evaluation of the translational disconnect between experimental and clinical sepsis can be made. Methods THP-1 (human) cells and RAW 264.7 (murine) cells were dosed with concentrations of LPS (human, 1 pg/mL to 100 ng/mL; murine, 30 pg/mL to 1,000 ng/mL) and nuclear actin and p65 were immunoblotted to measure changes in nuclear density. In vivo, C57BL/6 mice received either IP injection of stool suspension (5 µL/g) or LPS (25 mg/kg) or saline (1 mL/kg). Animals were culled at 6 hours and tissues were analyzed. Results An increase in basal p65:actin density in THP-1 cells (mean 0.214, standard error of the mean 0.024) was seen at doses as small as 0.1 ng/mL (0.519±0.064). In contrast to RAW 264.7 cells, basal increases (0.170±0.025) were only seen when a dose of 3 ng/mL (0.387±0.078) was used. Dose–response analysis of p65:actin ratio showed that THP-1 cells respond to lower doses of LPS than RAW 264.7 cells and lower doses produce a greater fold increase in the nuclear p65 density. Both in vivo models showed evidence of neutrophil (NL) recruitment into tissues (which was more intense after LPS treatment). IP stool inoculation resulted in an acute suppurative peritonitis and more substantial evidence of NL recruitment into adipose tissue and skeletal muscle

  9. Claudin 7 expression and localization in the normal murine mammary gland and murine mammary tumors

    Claudins, membrane-associated tetraspanin proteins, are normally associated with the tight junctions of epithelial cells where they confer a variety of permeability properties to the transepithelial barrier. One member of this family, claudin 7, has been shown to be expressed in the human mammary epithelium and some breast tumors. To set the stage for functional experiments on this molecule, we examined the developmental expression and localization of claudin 7 in the murine mammary epithelium and in a selection of murine mammary tumors. We used real-time polymerase chain reaction, in situ mRNA localization, and immunohistochemistry (IHC) to examine the expression and localization of claudin 7. Frozen sections were examined by digital confocal microscopy for colocalization with the tight-junction protein ZO1. Claudin 7 was expressed constitutively in the mammary epithelium at all developmental stages, and the ratio of its mRNA to that of keratin 19 was nearly constant through development. By IHC, claudin 7 was located in the basolateral part of the cell where it seemed to be localized to discrete vesicles. Scant colocalization with the tight-junction scaffolding protein ZO1 was observed. Similar results were obtained from IHC of the airway epithelium and some renal tubules; however, claudin 7 did partly colocalize with ZO1 in EPH4 cells, a normal murine mammary cell line, and in the epididymis. The molecule was localized in the cytoplasm of MMTV-neu and the transplantable murine tumor cell lines TM4, TM10, and TM40A, in which its ratio to cytokeratin was higher than in the normal mammary epithelium. Claudin 7 is expressed constitutively in the mammary epithelium at approximately equal levels throughout development as well as in the murine tumors examined. Although it is capable of localizing to tight junctions, in the epithelia of mammary gland, airway, and kidney it is mostly or entirely confined to punctate cytoplasmic structures, often near the basolateral

  10. Partial characterization of murine migration inhibitory factor (MIF).

    Kühner, A L; David, J R

    1976-01-01

    These studies describe the production of murine migration inhibitory factor (MIF)3 in sufficient quantities to allow its partial characterization by physiochemical and enzymatic methods. MIF was obtained from murine spleen cell cultures (C57BL/6 strain) stimulated with concanavalin A (Con A). Characterization of murine MIF was performed using Sephadex G-100 gel chromatography, isopycnic centrifugation in a CsCl density gradient, polyacrylamide disc electrophoresis, heat stability, and enzymatic treatment. MIF-containing and control fractions were assayed on normal C57BL/6 peritoneal exudate cells by using a microcapillary tube assay. Peak MIF activity was found in a Sephadex G-100 fraction containing molecules the size of albumin and slightly smaller, molecular weight 67,000 to 48,000. Murine MIF was stable to heating at 56 degrees C for 30 min but lost its activity at 80 degrees C for 30 min. Incubation of G-100 fractions containing MIF with water insoluble chymotrypsin destroyed the activity of MIF, indicating its protein nature. CsCl density gradient centrifugation revealed that murine MIF had a buoyand density greater than protein, consistent with its being a glycoprotein. Further, when subjected to disc electrophoresis on polyacylamide gels, murine MIF migrated in a region cathodal to albumin. Thus, mitogen stimulation of murine spleen cells produced MIF in quantities which allowed its partial characterization and purification, and its comparison with human and guinea pig MIF; this makes it feasible to analyze the role of murine MIF in cellular immunity and in its relationship to lymphocyte mediators which regulate humoral immune responses. PMID:1107423

  11. Human recombinant erythropoietin promotes differentiation of murine megakaryocytes in vitro.

    Ishibashi, T.; Koziol, J A; Burstein, S A

    1987-01-01

    To determine if erythropoietin affects megakaryocytopoiesis, we measured acetylcholinesterase (AchE) activity, a marker of the murine megakaryocytic lineage, after the addition of human recombinant erythropoietin to serumless murine bone marrow cultures. Erythropoietin increased AchE activity substantially. Moreover, when the hormone was added to serumless cultures of 426 isolated single megakaryocytes derived from megakaryocytic colonies, erythropoietin induced a significant increase in the ...

  12. Dynamic Determination of Oxygenation and Lung Compliance in Murine Pneumonectomy

    Gibney, Barry; Lee, Grace S; Houdek, Jan; Lin, Miao; Miele, Lino; Chamoto, Kenji; Konerding, Moritz A; Tsuda, Akira; Mentzer, Steven J.

    2011-01-01

    Thoracic surgical procedures in mice have been applied to a wide range of investigations, but little is known about the murine physiologic response to pulmonary surgery. Using continuous arterial oximetry monitoring and the FlexiVent murine ventilator, we investigated the effect of anesthesia and pneumonectomy on mouse oxygen saturation and lung mechanics. Sedation resulted in a dose-dependent decline of oxygen saturation that ranged from 55–82%. Oxygen saturation was restored by mechanical v...

  13. Surface Contaminants Inhibit Osseointegration in a Novel Murine Model

    Bonsignore, Lindsay A.; Colbrunn, Robb W.; Tatro, Joscelyn M.; Messerschmitt, Patrick J.; Hernandez, Christopher J.; Goldberg, Victor M.; Stewart, Matthew C.; Greenfield, Edward M.

    2011-01-01

    Surface contaminants, such as bacterial debris and manufacturing residues, may remain on orthopaedic implants after sterilization procedures and affect osseointegration. The goals of this study were to develop a murine model of osseointegration in order to determine whether removing surface contaminants enhances osseointegration. To develop the murine model, titanium alloy implants were implanted into a unicortical pilot hole in the mid-diaphysis of the femur and osseointegration was measured...

  14. microRNA-222 modulates liver fibrosis in a murine model of biliary atresia

    Shen, Wen-jun; Dong, Rui; Chen, Gong, E-mail: chengongzlp@hotmail.com; Zheng, Shan

    2014-03-28

    Highlights: • The RRV infected group showed cholestasis, retardation and extrahepatic biliary atresia. • miR-222 was highly expressed, and PPP2R2A was inhibited in the murine biliary atresia model. • miR-222 profoundly modulated the process of fibrosis in the murine biliary atresia model. • miR-222 might represent a potential target for improving biliary atresia prognosis. - Abstract: microRNA-222 (miR-222) has been shown to initiate the activation of hepatic stellate cells, which plays an important role in the pathogenesis of liver fibrosis. The aim of our study was to evaluate the role of miR-22 in a mouse model of biliary atresia (BA) induced by Rhesus Rotavirus (RRV) infection. New-born Balb/c mice were randomized into control and RRV infected groups. The extrahepatic bile ducts were evaluated. The experimental group was divided into BA group and negative group based on histology. The expression of miR-222, protein phosphatase 2 regulatory subunit B alpha (PPP2R2A), proliferating cell nuclear antigen (PCNA) and phospho-Akt were detected. We found that the experimental group showed signs of cholestasis, retardation and extrahepatic biliary atresia. No abnormalities were found in the control group. In the BA group, miR-222, PCNA and Akt were highly expressed, and PPP2R2A expression was significantly inhibited. Our findings suggest that miR-222 profoundly modulated the process of fibrosis in the murine BA model, which might represent a potential target for improving BA prognosis.

  15. microRNA-222 modulates liver fibrosis in a murine model of biliary atresia

    Highlights: • The RRV infected group showed cholestasis, retardation and extrahepatic biliary atresia. • miR-222 was highly expressed, and PPP2R2A was inhibited in the murine biliary atresia model. • miR-222 profoundly modulated the process of fibrosis in the murine biliary atresia model. • miR-222 might represent a potential target for improving biliary atresia prognosis. - Abstract: microRNA-222 (miR-222) has been shown to initiate the activation of hepatic stellate cells, which plays an important role in the pathogenesis of liver fibrosis. The aim of our study was to evaluate the role of miR-22 in a mouse model of biliary atresia (BA) induced by Rhesus Rotavirus (RRV) infection. New-born Balb/c mice were randomized into control and RRV infected groups. The extrahepatic bile ducts were evaluated. The experimental group was divided into BA group and negative group based on histology. The expression of miR-222, protein phosphatase 2 regulatory subunit B alpha (PPP2R2A), proliferating cell nuclear antigen (PCNA) and phospho-Akt were detected. We found that the experimental group showed signs of cholestasis, retardation and extrahepatic biliary atresia. No abnormalities were found in the control group. In the BA group, miR-222, PCNA and Akt were highly expressed, and PPP2R2A expression was significantly inhibited. Our findings suggest that miR-222 profoundly modulated the process of fibrosis in the murine BA model, which might represent a potential target for improving BA prognosis

  16. Experimental study of hematopoietic stem cell transplantation for the treatment of murine systemic lupus erythematosus%混合造血干细胞移植治疗小鼠系统性红斑狼疮的实验研究

    倪丽; 刘冠贤; 董光富; 石咏军; 崔晓

    2010-01-01

    Objective To study the effect of mixed purified autogenic and allogeneic hematopoietic stem cell transplantation for the treatment of systemic lupus erythematosus. Methods Thirty-six MRL/lpr mice were randomly divided into the control group, the study group,the mixed group ( the ratio of autogenic to hematopoietic stem cells, mixed in different proportions were infused intravenously after 60Co irradiation. The study group were treated with daily intraperitoneal infusion of dexamethasone 1 mg·kg-1·d-1, while the control group were treated with intraperitoneal infusion of equivalent volume of saline daily. The changes of serum creatinine level, the urine protein excretion of the mice and blood WBC count were compared. Repeat measures ANOVA was used for data analysis. ELISA was used for anti-nuclear antibody detection Light microscopy, electronic micros-copy, immunofluorescence were applied to detect the pathological changes in renal tissue. Results Serum creatinine and urine protein excretion levels increased with time in the ontrol group, while those of the transplant group and the study group decreased. The reduction in mixed transplantation group and the study group was more evident compared with that of the allogeneic group. The difference was statistically significant (P<0.05), but there was no significant difference between the mixed transplantation groups and the study group (P>0.05). The histopathologic damage was most serious in the control group as pathological injury score of most mice were in grade 3 or 4. The majority of the histopathologic damage of the allogeneic group was in grade 2. Most f pathological damage of the study drug group and the mixed transplantation group were grade 1 or 2. Conclusion Mixed hematoopoietic stem cell transplantation for the treatment of murine systemic lupus erythematosus can effectively correct heavy proteinuria in murine systemic lupus erythematosus so improve the renal damage. It is a safe and effectively way to

  17. Human glioblastoma-associated microglia/monocytes express a distinct RNA profile compared to human control and murine samples.

    Szulzewsky, Frank; Arora, Sonali; de Witte, Lot; Ulas, Thomas; Markovic, Darko; Schultze, Joachim L; Holland, Eric C; Synowitz, Michael; Wolf, Susanne A; Kettenmann, Helmut

    2016-08-01

    Glioblastoma (GBM) is the most aggressive brain tumor in adults. It is strongly infiltrated by microglia and peripheral monocytes that support tumor growth. In the present study we used RNA sequencing to compare the expression profile of CD11b(+) human glioblastoma-associated microglia/monocytes (hGAMs) to CD11b(+) microglia isolated from non-tumor samples. Hierarchical clustering and principal component analysis showed a clear separation of the two sample groups and we identified 334 significantly regulated genes in hGAMs. In comparison to human control microglia hGAMs upregulated genes associated with mitotic cell cycle, cell migration, cell adhesion, and extracellular matrix organization. We validated the expression of several genes associated with extracellular matrix organization in samples of human control microglia, hGAMs, and the hGAMs-depleted fraction via qPCR. The comparison to murine GAMs (mGAMs) showed that both cell populations share a significant fraction of upregulated transcripts compared with their respective controls. These genes were mostly related to mitotic cell cycle. However, in contrast to murine cells, human GAMs did not upregulate genes associated to immune activation. Comparison of human and murine GAMs expression data to several data sets of in vitro-activated human macrophages and murine microglia showed that, in contrast to mGAMs, hGAMs share a smaller overlap to these data sets in general and in particular to cells activated by proinflammatory stimulation with LPS + INFγ or TNFα. Our findings provide new insights into the biology of human glioblastoma-associated microglia/monocytes and give detailed information about the validity of murine experimental models. GLIA 2016 GLIA 2016;64:1416-1436. PMID:27312099

  18. Research on Growth Behavior of Embryos for Bovine and Murine on Primary Murine Embryos Fibroblast Cell Feeder Layer

    AN Li-long; XIAO Mei; FENG Xiu-Liang; DOU Zhong-ying; QIU Huai; YANG Qi; LEI An-min; YANG Chun-rong; GAO Zhi-min

    2002-01-01

    The difference in growth behavior between bovine embryos and murine embryos was studied on PMEF(primary murine embryos fibroblast)feeder layer. The results showed as follows: With embryos having attached, bovine embryonic trophoblast formed a transparent membranous structure covering on inner cell mass (ICM), however, murine embryonic trophoblast formed disc structure. Bovine embryos formed four kinds of ICM colonies with different morphology including the mass-like, the net-like, the stream-like and the mixture-like colonies. Compared with Murine ICM, the bovine ICM grew more fast. So, the bovine ICM was passaged at first after a culture of approximately 5 - 6 days in vitro, but murine ICM was passaged at first after an attachment of 3 - 4 days on PMEF feeder layer. The mixture colonies of bovine ICM differentiated very early, while the others differentiated very late. Most ICM-like mass of Bovine grew in a defined spot, but bovine ICMs like stream and ICMs like net proliferated fast and dispersed quickly. We found that the single blastomeres derived from late bovine morula and late murine morula formed sub-blastophere; moreover, the bovine ICM cell would differentiate rapidly if the trophoblast was removed.

  19. Tetrapyrrole binding affinity of the murine and human p22HBP heme-binding proteins.

    Micaelo, Nuno M; Macedo, Anjos L; Goodfellow, Brian J; Félix, Vítor

    2010-11-01

    We present the first systematic molecular modeling study of the binding properties of murine (mHBP) and human (hHBP) p22HBP protein (heme-binding protein) with four tetrapyrrole ring systems belonging to the heme biosynthetic pathway: iron protoporphyrin IX (HEMIN), protoporphyrin IX (PPIX), coproporphyrin III (CPIII), coproporphyrin I (CPI). The relative binding affinities predicted by our computational study were found to be similar to those observed experimentally, providing a first rational structural analysis of the molecular recognition mechanism, by p22HBP, toward a number of different tetrapyrrole ligands. To probe the structure of these p22HBP protein complexes, docking, molecular dynamics and MM-PBSA methodologies supported by experimental NMR ring current shift data have been employed. The tetrapyrroles studied were found to bind murine p22HBP with the following binding affinity order: HEMIN> PPIX> CPIII> CPI, which ranged from -22.2 to -6.1 kcal/mol. In general, the protein-tetrapyrrole complexes are stabilized by non-bonded interactions between the tetrapyrrole propionate groups and basic residues of the protein, and by the preferential solvation of the complex compared to the unbound components. PMID:20800521

  20. Remodeling of alveolar septa after murine pneumonectomy.

    Ysasi, Alexandra B; Wagner, Willi L; Bennett, Robert D; Ackermann, Maximilian; Valenzuela, Cristian D; Belle, Janeil; Tsuda, Akira; Konerding, Moritz A; Mentzer, Steven J

    2015-06-15

    In most mammals, removing one lung (pneumonectomy) results in the compensatory growth of the remaining lung. In mice, stereological observations have demonstrated an increase in the number of mature alveoli; however, anatomic evidence of the early phases of alveolar growth has remained elusive. To identify changes in the lung microstructure associated with neoalveolarization, we used tissue histology, electron microscopy, and synchrotron imaging to examine the configuration of the alveolar duct after murine pneumonectomy. Systematic histological examination of the cardiac lobe demonstrated no change in the relative frequency of dihedral angle components (Ends, Bends, and Junctions) (P > 0.05), but a significant decrease in the length of a subset of septal ends ("E"). Septal retraction, observed in 20-30% of the alveolar ducts, was maximal on day 3 after pneumonectomy (P alveolar duct diameter ratio (Dout:Din) was significantly lower 3 days after pneumonectomy compared to all controls except for the detergent-treated lung (P surface tension within the alveolar duct, resulting in a new equilibrium at a higher total energy and lower surface area. The spatial and temporal association of these microstructural changes with postpneumonectomy lung growth suggests that these changes represent an early phase of alveolar duct remodeling. PMID:26078396

  1. ESCRT Requirements for Murine Leukemia Virus Release

    Bartusch, Christina; Prange, Reinhild

    2016-01-01

    The Murine Leukemia Virus (MLV) is a gammaretrovirus that hijack host components of the endosomal sorting complex required for transport (ESCRT) for budding. To determine the minimal requirements for ESCRT factors in MLV viral and viral-like particles (VLP) release, an siRNA knockdown screen of ESCRT(-associated) proteins was performed in MLV-producing human cells. We found that MLV VLPs and virions primarily engage the ESCRT-I factor Tsg101 and marginally the ESCRT-associated adaptors Nedd4-1 and Alix to enter the ESCRT pathway. Conversely, the inactivation of ESCRT-II had no impact on VLP and virion egress. By analyzing the effects of individual ESCRT-III knockdowns, VLP and virion release was profoundly inhibited in CHMP2A- and CHMP4B-knockdown cells. In contrast, neither the CHMP2B and CHMP4A isoforms nor CHMP3, CHMP5, and CHMP6 were found to be essential. In case of CHMP1, we unexpectedly observed that the CHMP1A isoform was specifically required for virus budding, but dispensable for VLP release. Hence, MLV utilizes only a subset of ESCRT factors, and viral and viral-like particles differ in ESCRT-III factor requirements. PMID:27096867

  2. Glycosaminoglycan interactions in murine gammaherpesvirus-68 infection.

    Laurent Gillet

    Full Text Available Glycosaminoglycans (GAGs commonly participate in herpesvirus entry. They are thought to provide a reversible attachment to cells that promotes subsequent receptor binding. Murine gamma-herpesvirus-68 (MHV-68 infection of fibroblasts and epithelial cells is highly GAG-dependent. This is a function of the viral gp150, in that gp150-deficient mutants are much less GAG-dependent than wild-type. Here we show that the major MHV-68 GAG-binding protein is not gp150 but gp70, a product of ORF4. Surprisingly, ORF4-deficient MHV-68 showed normal cell binding and was more sensitive than wild-type to inhibition by soluble heparin rather than less. Thus, the most obvious viral GAG interaction made little direct contribution to infection. Indeed, a large fraction of the virion gp70 had its GAG-binding domain removed by post-translational cleavage. ORF4 may therefore act mainly to absorb soluble GAGs and prevent them from engaging gp150 prematurely. In contrast to gp70, gp150 bound poorly to GAGs, implying that it provides little in the way of adhesion. We hypothesize that it acts instead as a GAG-sensitive switch that selectively activates MHV-68 entry at cell surfaces.

  3. Structural and biochemical characterization of the inhibitor complexes of xenotropic murine leukemia virus-related virus protease

    Li, Mi; Gustchina, Alla; Matúz, Krisztina; Tözsér, Jozsef; Namwong, Sirilak; Goldfarb, Nathan E.; Dunn, Ben M.; Wlodawer, Alexander (Debrecen); (NCI); (Florida); (Suan Sunandha)

    2012-10-23

    Interactions between the protease (PR) encoded by the xenotropic murine leukemia virus-related virus and a number of potential inhibitors have been investigated by biochemical and structural techniques. It was observed that several inhibitors used clinically against HIV PR exhibit nanomolar or even subnanomolar values of K{sub i}, depending on the exact experimental conditions. Both TL-3, a universal inhibitor of retroviral PRs, and some inhibitors originally shown to inhibit plasmepsins were also quite potent, whereas inhibition by pepstatin A was considerably weaker. Crystal structures of the complexes of xenotropic murine leukemia virus-related virus PR with TL-3, amprenavir and pepstatin A were solved at high resolution and compared with the structures of complexes of these inhibitors with other retropepsins. Whereas TL-3 and amprenavir bound in a predictable manner, spanning the substrate-binding site of the enzyme, two molecules of pepstatin A bound simultaneously in an unprecedented manner, leaving the catalytic water molecule in place.

  4. Demonstration of a cell wall antigen cross-reacting with cryptococcal polysaccharide in experimental disseminated trichosporonosis.

    Melcher, G P; Reed, K D; Rinaldi, M. G.; Lee, J. W.; Pizzo, P A; Walsh, T. J.

    1991-01-01

    Patients with disseminated infections caused by Trichosporon beigelii have a circulating antigen that cross-reacts with the polysaccharide capsule of Cryptococcus neoformans. We studied the localization of this antigen by immunoelectron microscopy in a rabbit model of experimental disseminated trichosporonosis. Deparaffinized lung sections were examined by using a murine monoclonal anti-cryptococcal polysaccharide antibody and colloidal gold particles coated with goat antibody to murine immun...

  5. Forced recombination of psi-modified murine leukaemia virus-based vectors with murine leukaemia-like and VL30 murine endogenous retroviruses

    Mikkelsen, J G; Lund, Anders Henrik; Duch, M;

    1999-01-01

    -impaired Akv-MLV-derived vectors, we here examine putative genetic interactions between vector RNAs and copackaged endogenous retroviral RNAs of the murine leukaemia virus (MLV) and VL30 retroelement families. We show (i) that MLV recombination is not blocked by nonhomology within the 5' untranslated region...

  6. Non- invasive in vivo analysis of a murine aortic graft using high resolution ultrasound microimaging

    Introduction: As yet, murine aortic grafts have merely been monitored histopathologically. The aim of our study was to examine how these grafts can be monitored in vivo and non-invasively by using high-resolution ultrasound microimaging to evaluate function and morphology. A further aim was to prove if this in vivo monitoring can be correlated to immunohistological data that indicates graft integrity. Methods: Murine infrarenal aortic isografts were orthotopically transplanted into 14 female mice (C57BL/6-Background) whereas a group of sham-operated animals (n = 10) served as controls. To assess the graft morphology and hemodynamics, we examined the mice over a post-operative period of 8 weeks with a sophisticated ultrasound system (Vevo 770, Visual Sonics). Results: The non-invasive graft monitoring was feasible in all transplanted mice. We could demonstrate a regular post-transplant graft function and morphology, such as anterior/posterior wall displacement and wall thickness. Mild alterations of anterior wall motion dynamics could only be observed at the site of distal graft anastomosis (8 weeks after grafting (transplant vs. sham mice: 0.02 mm ± 0.01 vs. 0.03 mm ± 0.01, p < 0.05). However, the integrity of the entire graft wall could be confirmed by histopathological evaluation of the grafts. Conclusions: With regard to graft patency, function and morphology, high resolution ultrasound microimaging has proven to be a valuable tool for longitudinal, non-invasive, in vivo graft monitoring in this murine aortic transplantation model. Consequently, this experimental animal model provides an excellent basis for molecular and pharmacological studies using genetically engineered mice.

  7. Analysis of cardiomyocyte movement in the developing murine heart

    Hashimoto, Hisayuki [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan); Yuasa, Shinsuke, E-mail: yuasa@a8.keio.jp [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan); Tabata, Hidenori [Department of Anatomy, Keio University School of Medicine, Tokyo (Japan); Tohyama, Shugo; Seki, Tomohisa; Egashira, Toru; Hayashiji, Nozomi; Hattori, Fumiyuki; Kusumoto, Dai; Kunitomi, Akira; Takei, Makoto; Kashimura, Shin; Yozu, Gakuto; Shimojima, Masaya; Motoda, Chikaaki; Muraoka, Naoto [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan); Nakajima, Kazunori [Department of Anatomy, Keio University School of Medicine, Tokyo (Japan); Sakaue-Sawano, Asako; Miyawaki, Atsushi [Life Function and Dynamics, ERATO, JST, 2-1 Hirosawa, Wako-city, Saitama 351-0198 (Japan); Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-city, Saitama 351-0198 (Japan); Fukuda, Keiichi [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan)

    2015-09-04

    The precise assemblage of several types of cardiac precursors controls heart organogenesis. The cardiac precursors show dynamic movement during early development and then form the complicated heart structure. However, cardiomyocyte movements inside the newly organized mammalian heart remain unclear. We previously established the method of ex vivo time-lapse imaging of the murine heart to study cardiomyocyte behavior by using the Fucci (fluorescent ubiquitination-based cell cycle indicator) system, which can effectively label individual G1, S/G2/M, and G1/S-transition phase nuclei in living cardiomyocytes as red, green, and yellow, respectively. Global analysis of gene expression in Fucci green positive ventricular cardiomyocytes confirmed that cell cycle regulatory genes expressed in G1/S, S, G2/M, and M phase transitions were upregulated. Interestingly, pathway analysis revealed that many genes related to the cell cycle were significantly upregulated in the Fucci green positive ventricular cardiomyocytes, while only a small number of genes related to cell motility were upregulated. Time-lapse imaging showed that murine proliferating cardiomyocytes did not exhibit dynamic movement inside the heart, but stayed on site after entering the cell cycle. - Highlights: • We directly visualized cardiomyocyte movement inside the developing murine heart. • Cell cycle related genes were upregulated in the proliferating cardiomyocytes. • Time-lapse imaging revealed that proliferating murine cardiomyocytes stayed in place. • Murine ventricular cardiomyocytes proliferate on site during development.

  8. Analysis of cardiomyocyte movement in the developing murine heart

    The precise assemblage of several types of cardiac precursors controls heart organogenesis. The cardiac precursors show dynamic movement during early development and then form the complicated heart structure. However, cardiomyocyte movements inside the newly organized mammalian heart remain unclear. We previously established the method of ex vivo time-lapse imaging of the murine heart to study cardiomyocyte behavior by using the Fucci (fluorescent ubiquitination-based cell cycle indicator) system, which can effectively label individual G1, S/G2/M, and G1/S-transition phase nuclei in living cardiomyocytes as red, green, and yellow, respectively. Global analysis of gene expression in Fucci green positive ventricular cardiomyocytes confirmed that cell cycle regulatory genes expressed in G1/S, S, G2/M, and M phase transitions were upregulated. Interestingly, pathway analysis revealed that many genes related to the cell cycle were significantly upregulated in the Fucci green positive ventricular cardiomyocytes, while only a small number of genes related to cell motility were upregulated. Time-lapse imaging showed that murine proliferating cardiomyocytes did not exhibit dynamic movement inside the heart, but stayed on site after entering the cell cycle. - Highlights: • We directly visualized cardiomyocyte movement inside the developing murine heart. • Cell cycle related genes were upregulated in the proliferating cardiomyocytes. • Time-lapse imaging revealed that proliferating murine cardiomyocytes stayed in place. • Murine ventricular cardiomyocytes proliferate on site during development

  9. Murine Typhus: An Important Consideration for the Nonspecific Febrile Illness

    Gurjot Basra

    2012-01-01

    Full Text Available Murine typhus is a widely distributed flea-borne infection caused by Rickettsia typhi. Symptoms of murine typhus are nonspecific and mimic a variety of other infectious diseases. We herein report a case of murine typhus in an area where the broad use of DDT in the mid-20th century has now made it a rare disease. The patient described presented with headache, fever, and a faint macular rash. Initial laboratory studies revealed a slight transaminase elevation. Further questioning revealed exposure to opossums, prompting the consideration of murine typhus as a diagnosis. Although typhus group antibodies were not present during the patient’s acute illness, empiric therapy with doxycycline was initiated, and the patient defervesced. One month after convalescence, the patient returned to clinic with serum that contained typhus group antibodies with an IgG titer of 1 : 1024. Murine typhus is an important consideration during the workup of a patient with a nonspecific febrile illness. Exposure to reservoir hosts and the flea vector place humans at risk for this disease. Clinician recognition of this entity is required for diagnosis and effective therapy.

  10. Evidence for intermittent radiobiological hypoxia in experimental tumour systems

    This paper describes flow and static fluorescence cytometry techniques to visualize and quantitate acute radiobiological hypoxia resulting from transient fluctuation in tumour blood flow in experimental tumour systems. The application of these techniques in two murine tumour systems provides evidence that such hypoxia exists and reduces the effectiveness of single doses of radiation. Possible mechanisms for and implications of these findings are discussed. (author)

  11. A rapid murine coma and behavior scale for quantitative assessment of murine cerebral malaria.

    Ryan W Carroll

    Full Text Available BACKGROUND: Cerebral malaria (CM is a neurological syndrome that includes coma and seizures following malaria parasite infection. The pathophysiology is not fully understood and cannot be accounted for by infection alone: patients still succumb to CM, even if the underlying parasite infection has resolved. To that effect, there is no known adjuvant therapy for CM. Current murine CM (MCM models do not allow for rapid clinical identification of affected animals following infection. An animal model that more closely mimics the clinical features of human CM would be helpful in elucidating potential mechanisms of disease pathogenesis and evaluating new adjuvant therapies. METHODOLOGY/PRINCIPAL FINDINGS: A quantitative, rapid murine coma and behavior scale (RMCBS comprised of 10 parameters was developed to assess MCM manifested in C57BL/6 mice infected with Plasmodium berghei ANKA (PbA. Using this method a single mouse can be completely assessed within 3 minutes. The RMCBS enables the operator to follow the evolution of the clinical syndrome, validated here by correlations with intracerebral hemorrhages. It provides a tool by which subjects can be identified as symptomatic prior to the initiation of trial treatment. CONCLUSIONS/SIGNIFICANCE: Since the RMCBS enables an operator to rapidly follow the course of disease, label a subject as affected or not, and correlate the level of illness with neuropathologic injury, it can ultimately be used to guide the initiation of treatment after the onset of cerebral disease (thus emulating the situation in the field. The RMCBS is a tool by which an adjuvant therapy can be objectively assessed.

  12. Cell therapy in the treatment of bronchiolitis obliterans in a murine model

    Julio de Oliveira Espinel

    2015-06-01

    Full Text Available OBJECTIVE: To evaluate the importance of stem cells derived from adipose tissue in reducing graft inflammation in a murine model of allogeneic heterotopic tracheal transplant.METHODS: We performed a heterotopic tracheal allografting in dorsal subcutaneous pouch and systemically injected 5x105 mesenchymal stem cells derived from adipose tissue. The animals were divided into two groups according to the time of sacrifice: T7 and T21. We also carried out histological analysis and digital morphometry.RESULTS: The T7 animals treated with cell therapy had median obstructed graft area of 0 versus 0.54 of controls (p = 0.635. The treated T21 subjects had median obstructed graft area of 0.25 versus 0 in controls (p = 0.041.CONCLUSION: The systemically injected cell therapy in experimental murine model of bronchiolitis obliterans did not reduce the severity of the allograft inflammation in a statistically significant way in seven days; Conversely, in 21 days, it increased the allograft inflammatory process.

  13. Femur Window Chamber Model for In Vivo Cell Tracking in the Murine Bone Marrow.

    Chen, Yonghong; Maeda, Azusa; Bu, Jiachuan; DaCosta, Ralph

    2016-01-01

    Bone marrow is a complex organ that contains various hematopoietic and non-hematopoietic cells. These cells are involved in many biological processes, including hematopoiesis, immune regulation and tumor regulation. Commonly used methods for understanding cellular actions in the bone marrow, such as histology and blood counts, provide static information rather than capturing the dynamic action of multiple cellular components in vivo. To complement the standard methods, a window chamber (WC)-based model was developed to enable serial in vivo imaging of cells and structures in the murine bone marrow. This protocol describes a surgical procedure for installing the WC in the femur, in order to facilitate long-term optical access to the femoral bone marrow. In particular, to demonstrate its experimental utility, this WC approach was used to image and track neutrophils within the vascular network of the femur, thereby providing a novel method to visualize and quantify immune cell trafficking and regulation in the bone marrow. This method can be applied to study various biological processes in the murine bone marrow, such as hematopoiesis, stem cell transplantation, and immune responses in pathological conditions, including cancer. PMID:27500928

  14. Dendritic cell-based vaccination in cancer: therapeutic implications emerging from murine models

    Soledad eMac Keon

    2015-05-01

    Full Text Available Dendritic cells (DCs play a pivotal role in the orchestration of immune responses, and are thus key targets in cancer vaccine design. Since the 2010 FDA approval of the first cancer DC-based vaccine (Sipuleucel T there has been a surge of interest in exploiting these cells as a therapeutic option for the treatment of tumors of diverse origin. In spite of the encouraging results obtained in the clinic, many elements of DC-based vaccination strategies need to be optimized. In this context, the use of experimental cancer models can help direct efforts towards an effective vaccine design. This paper reviews recent findings in murine models regarding the antitumoral mechanisms of DC-based vaccination, covering issues related to antigen sources, the use of adjuvants and maturing agents, and the role of DC subsets and their interaction in the initiation of antitumoral immune responses. The summary of such diverse aspects will highlight advantages and drawbacks in the use of murine models, and contribute to the design of successful DC-based translational approaches for cancer treatment.

  15. Mutational dynamics of murine angiogenin duplicates

    Fares Mario A

    2010-10-01

    Full Text Available Abstract Background Angiogenin (Ang is a protein involved in angiogenesis by inducing the formation of blood vessels. The biomedical importance of this protein has come from findings linking mutations in Ang to cancer progression and neurodegenerative diseases. These findings highlight the evolutionary constrain on Ang amino acid sequence. However, previous studies comparing human Angiogenin with homologs from other phylogenetically related organisms have led to the conclusion that Ang presents a striking variability. Whether this variability has an adaptive value per se remains elusive. Understanding why many functional Ang paralogs have been preserved in mouse and rat and identifying functional divergence mutations at these copies may explain the relationship between mutations and function. In spite of the importance of testing this hypothesis from the evolutionarily and biomedical perspectives, this remains yet unaccomplished. Here we test the main mutational dynamics driving the evolution and function of Ang paralogs in mammals. Results We analysed the phylogenetic asymmetries between the different Ang gene copies in mouse and rat in the context of vertebrate Ang phylogeny. This analysis shows strong evidence in support of accelerated evolution in some Ang murine copies (mAng. This acceleration is not due to non-functionalisation because constraints on amino acid replacements remain strong. We identify many of the amino acid sites involved in signal localization and nucleotide binding by Ang to have evolved under diversifying selection. Compensatory effects of many of the mutations at these paralogs and their key structural location in or nearby important functional regions support a possible functional shift (functional divergence in many Ang copies. Similarities between 3D-structural models for mAng copies suggest that their divergence is mainly functional. Conclusions We identify the main evolutionary dynamics shaping the variability of

  16. Nanoelectroablation therapy for murine basal cell carcinoma

    Highlights: ► Nanoelectroablation is a new, non-thermal therapy that triggers apoptosis in tumors. ► Low energy, ultrashort, high voltage pulses ablate the tumor with little or no scar. ► Nanoelectroablation eliminates 99.8% of the BCC but may leave a few remnants behind. ► Pilot clinical trials on human BCCs are ongoing and leave no remnants in most cases. -- Abstract: When skin tumors are exposed to non-thermal, low energy, nanosecond pulsed electric fields (nsPEF), apoptosis is initiated both in vitro and in vivo. This nanoelectroablation therapy has already been proven effective in treating subdermal murine allograft tumors. We wanted to determine if this therapy would be equally effective in the treatment of autochthonous BCC tumors in Ptch1+/−K14-Cre-ER p53 fl/fl mice. These tumors are similar to human BCCs in histology and in response to drug therapy . We have treated 27 BCCs across 8 mice with either 300 pulses of 300 ns duration or 2700 pulses of 100 ns duration, all at 30 kV/cm and 5–7 pulses per second. Every nsPEF-treated BCC began to shrink within a day after treatment and their initial mean volume of 36 ± 5 (SEM) mm3 shrunk by 76 ± 3% over the ensuing two weeks. After four weeks, they were 99.8% ablated if the size of the treatment electrode matched the tumor size. If the tumor was larger than the 4 mm wide electrode, multiple treatments were needed for complete ablation. Treated tumors were harvested for histological analysis at various times after treatment and exhibited apoptosis markers. Specifically, pyknosis of nuclei was evident as soon as 2 days after nsPEF treatment, and DNA fragmentation as detected via TUNEL staining was also evident post treatment. Nanoelectroablation is effective in triggering apoptosis and remission of radiation-induced BCCs with a single 6 min-long treatment of 2700 pulses.

  17. Nanoelectroablation therapy for murine basal cell carcinoma

    Nuccitelli, Richard, E-mail: rich@bioelectromed.com [BioElectroMed Corp., 849 Mitten Rd., Suite 104, Burlingame, CA 94010 (United States); Tran, Kevin; Athos, Brian; Kreis, Mark; Nuccitelli, Pamela [BioElectroMed Corp., 849 Mitten Rd., Suite 104, Burlingame, CA 94010 (United States); Chang, Kris S.; Epstein, Ervin H. [The Children' s Hospital Oakland Research Institute, Oakland, CA 94609 (United States); Tang, Jean Y. [The Children' s Hospital Oakland Research Institute, Oakland, CA 94609 (United States); Stanford University, Stanford, CA 94305 (United States)

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer Nanoelectroablation is a new, non-thermal therapy that triggers apoptosis in tumors. Black-Right-Pointing-Pointer Low energy, ultrashort, high voltage pulses ablate the tumor with little or no scar. Black-Right-Pointing-Pointer Nanoelectroablation eliminates 99.8% of the BCC but may leave a few remnants behind. Black-Right-Pointing-Pointer Pilot clinical trials on human BCCs are ongoing and leave no remnants in most cases. -- Abstract: When skin tumors are exposed to non-thermal, low energy, nanosecond pulsed electric fields (nsPEF), apoptosis is initiated both in vitro and in vivo. This nanoelectroablation therapy has already been proven effective in treating subdermal murine allograft tumors. We wanted to determine if this therapy would be equally effective in the treatment of autochthonous BCC tumors in Ptch1{sup +/-}K14-Cre-ER p53 fl/fl mice. These tumors are similar to human BCCs in histology and in response to drug therapy . We have treated 27 BCCs across 8 mice with either 300 pulses of 300 ns duration or 2700 pulses of 100 ns duration, all at 30 kV/cm and 5-7 pulses per second. Every nsPEF-treated BCC began to shrink within a day after treatment and their initial mean volume of 36 {+-} 5 (SEM) mm{sup 3} shrunk by 76 {+-} 3% over the ensuing two weeks. After four weeks, they were 99.8% ablated if the size of the treatment electrode matched the tumor size. If the tumor was larger than the 4 mm wide electrode, multiple treatments were needed for complete ablation. Treated tumors were harvested for histological analysis at various times after treatment and exhibited apoptosis markers. Specifically, pyknosis of nuclei was evident as soon as 2 days after nsPEF treatment, and DNA fragmentation as detected via TUNEL staining was also evident post treatment. Nanoelectroablation is effective in triggering apoptosis and remission of radiation-induced BCCs with a single 6 min-long treatment of 2700 pulses.

  18. Chemokine (C-C motif ligand 2 mediates direct and indirect fibrotic responses in human and murine cultured fibrocytes

    Ekert Jason E

    2011-10-01

    Full Text Available Abstract Background Fibrocytes are a population of circulating bone-marrow-derived cells that express surface markers for leukocytes and mesenchymal cells, and are capable of differentiating into myofibroblasts. They have been observed at sites of active fibrosis and increased circulating numbers correlate with mortality in idiopathic pulmonary fibrosis (IPF. Inhibition of chemokine (C-C motif receptor 2 (CCR2 during experimental models of lung fibrosis reduces lung collagen deposition, as well as reducing lung fibrocyte accumulation. The aim of the present study was to determine whether human and mouse fibrocytes express functional CCR2. Results Following optimized and identical human and murine fibrocyte isolation, both cell sources were shown to be positive for CCR2 by flow cytometry and this expression colocalized with collagen I and CD45. Human blood fibrocytes stimulated with the CCR2 ligand chemokine (C-C motif ligand 2 (CCL2, demonstrated increased proliferation (P P P Conclusions This study directly compares the functional responses of human and murine fibrocytes to CCR2 ligands, and following comparable isolation techniques. We have shown comparable biological effects, strengthening the translatability of the murine models to human disease with respect to targeting the CCR2 axis to ameliorate disease in IPF patients.

  19. Association of murine lupus and thymic full-length endogenous retroviral expression maps to a bone marrow stem cell

    Krieg, A.M.; Gourley, M.F.; Steinberg, A.D. (National Institutes of Health, Bethesda, MD (USA))

    1991-05-01

    Recent studies of thymic gene expression in murine lupus have demonstrated 8.4-kb (full-length size) modified polytropic (Mpmv) endogenous retroviral RNA. In contrast, normal control mouse strains do not produce detectable amounts of such RNA in their thymuses. Prior studies have attributed a defect in experimental tolerance in murine lupus to a bone marrow stem cell rather than to the thymic epithelium; in contrast, infectious retroviral expression has been associated with the thymic epithelium, rather than with the bone marrow stem cell. The present study was designed to determine whether the abnormal Mpmv expression associated with murine lupus mapped to thymic epithelium or to a marrow precursor. Lethally irradiated control and lupus-prone mice were reconstituted with T cell depleted bone marrow; one month later their thymuses were studied for endogenous retroviral RNA and protein expression. Recipients of bone marrow from nonautoimmune donors expressed neither 8.4-kb Mpmv RNA nor surface MCF gp70 in their thymuses. In contrast, recipients of bone marrow from autoimmune NZB or BXSB donors expressed thymic 8.4-kb Mpmv RNA and mink cell focus-forming gp70. These studies demonstrate that lupus-associated 8.4-kb Mpmv endogenous retroviral expression is determined by bone marrow stem cells.

  20. Human Mesenchymal Stromal Cells Transplantation May Enhance or Inhibit 4T1 Murine Breast Adenocarcinoma through Different Approaches

    T. Jazedje

    2015-01-01

    Full Text Available The use of Mesenchymal Stromal Cells (MSCs aiming to treat cancer has shown very contradictory results. In an attempt to clarify the contradictory results reported in the literature and the possible role of human fallopian tube Mesenchymal Stromal Cells (htMSCs against breast cancer, the aim of this study was to evaluate the clinical effect of htMSCs in murine mammary adenocarcinoma using two different approaches: (1 coinjections of htMSCs and 4T1 murine tumor cell lineage and (2 injections of htMSCs in mice at the initial stage of mammary adenocarcinoma development. Coinjected animals had a more severe course of the disease and a reduced survival, while tumor-bearing animals treated with 2 intraperitoneal injections of 106 htMSCs showed significantly reduced tumor growth and increased lifespan as compared with control animals. Coculture of htMSCs and 4T1 tumor cells revealed an increase in IL-8 and MCP-1 and decreased VEGF production. For the first time, we show that MSCs isolated from a single source and donor when injected in the same animal model and tumor can lead to opposite results depending on the experimental protocol. Also, our results demonstrated that htMSCs can have an inhibitory effect on the development of murine mammary adenocarcinoma.

  1. Association of murine lupus and thymic full-length endogenous retroviral expression maps to a bone marrow stem cell

    Recent studies of thymic gene expression in murine lupus have demonstrated 8.4-kb (full-length size) modified polytropic (Mpmv) endogenous retroviral RNA. In contrast, normal control mouse strains do not produce detectable amounts of such RNA in their thymuses. Prior studies have attributed a defect in experimental tolerance in murine lupus to a bone marrow stem cell rather than to the thymic epithelium; in contrast, infectious retroviral expression has been associated with the thymic epithelium, rather than with the bone marrow stem cell. The present study was designed to determine whether the abnormal Mpmv expression associated with murine lupus mapped to thymic epithelium or to a marrow precursor. Lethally irradiated control and lupus-prone mice were reconstituted with T cell depleted bone marrow; one month later their thymuses were studied for endogenous retroviral RNA and protein expression. Recipients of bone marrow from nonautoimmune donors expressed neither 8.4-kb Mpmv RNA nor surface MCF gp70 in their thymuses. In contrast, recipients of bone marrow from autoimmune NZB or BXSB donors expressed thymic 8.4-kb Mpmv RNA and mink cell focus-forming gp70. These studies demonstrate that lupus-associated 8.4-kb Mpmv endogenous retroviral expression is determined by bone marrow stem cells

  2. Directed evolution and targeted mutagenesis to murinize listeria monocytogenes internalin A for enhanced infectivity in the murine oral infection model

    Hill Colin

    2010-12-01

    Full Text Available Abstract Background Internalin A (InlA is a critical virulence factor which mediates the initiation of Listeria monocytogenes infection by the oral route in permissive hosts. The interaction of InlA with the host cell ligand E-cadherin efficiently stimulates L. monocytogenes entry into human enterocytes, but has only a limited interaction with murine cells. Results We have created a surface display library of randomly mutated InlA in a non-invasive heterologous host Lactococcus lactis in order to create and screen novel variants of this invasion factor. After sequential passage through a murine cell line (CT-26, multiple clones with enhanced invasion characteristics were identified. Competitive index experiments were conducted in mice using selected mutations introduced into L. monocytogenes EGD-e background. A novel single amino acid change was identified which enhanced virulence by the oral route in the murine model and will form the basis of further engineering approaches. As a control a previously described EGD-InlAm murinized strain was also re-created as part of this study with minor modifications and designated EGD-e InlAm*. The strain was created using a procedure that minimizes the likelihood of secondary mutations and incorporates Listeria-optimized codons encoding the altered amino acids. L. monocytogenes EGD-e InlAm* yielded consistently higher level murine infections by the oral route when compared to EGD-e, but did not display the two-fold increased invasion into a human cell line that was previously described for the EGD-InlAm strain. Conclusions We have used both site-directed mutagenesis and directed evolution to create variants of InlA which may inform future structure-function analyses of this protein. During the course of the study we engineered a murinized strain of L. monocytogenes EGD-e which shows reproducibly higher infectivity in the intragastric murine infection model than the wild type, but does not display enhanced

  3. Directed evolution and targeted mutagenesis to murinize Listeria monocytogenes Internalin A for enhanced infectivity in the murine oral infection model

    Monk, Ian R

    2010-12-13

    Abstract Background Internalin A (InlA) is a critical virulence factor which mediates the initiation of Listeria monocytogenes infection by the oral route in permissive hosts. The interaction of InlA with the host cell ligand E-cadherin efficiently stimulates L. monocytogenes entry into human enterocytes, but has only a limited interaction with murine cells. Results We have created a surface display library of randomly mutated InlA in a non-invasive heterologous host Lactococcus lactis in order to create and screen novel variants of this invasion factor. After sequential passage through a murine cell line (CT-26), multiple clones with enhanced invasion characteristics were identified. Competitive index experiments were conducted in mice using selected mutations introduced into L. monocytogenes EGD-e background. A novel single amino acid change was identified which enhanced virulence by the oral route in the murine model and will form the basis of further engineering approaches. As a control a previously described EGD-InlAm murinized strain was also re-created as part of this study with minor modifications and designated EGD-e InlA m*. The strain was created using a procedure that minimizes the likelihood of secondary mutations and incorporates Listeria-optimized codons encoding the altered amino acids. L. monocytogenes EGD-e InlA m* yielded consistently higher level murine infections by the oral route when compared to EGD-e, but did not display the two-fold increased invasion into a human cell line that was previously described for the EGD-InlAm strain. Conclusions We have used both site-directed mutagenesis and directed evolution to create variants of InlA which may inform future structure-function analyses of this protein. During the course of the study we engineered a murinized strain of L. monocytogenes EGD-e which shows reproducibly higher infectivity in the intragastric murine infection model than the wild type, but does not display enhanced entry into human

  4. Immunohistochemical diagnosis of systemic bovine zygomycosis by murine monoclonal antibodies

    Jensen, H.E.; Aalbaek, B.; Lind, Peter;

    1996-01-01

    Murine monoclonal antibodies (Mabs) against water-soluble somatic antigens (WSSA) and the wall fraction (WF) from Rhizopus arrhizus (Rhizopus oryzae) were produced in vitro by fusion of splenocytes from immunized BALB/c mice with mouse myeloma X63-Ag 8.653 cells. Supernatants reacting only with h...

  5. Turnover of T cells in murine gammaherpesvirus 68-infected mice

    Hamilton-Easton, A M; Christensen, Jan Pravsgaard; Doherty, P C

    1999-01-01

    Respiratory challenge of C57BL/6 mice with murine gammaherpesvirus 68 induces proliferation of T lymphocytes early after infection, as evidenced by incorporation of the DNA precursor bromodeoxyuridine. Using pulse-chase analysis, splenic and peripheral blood activated T lymphocytes were found to ...

  6. Immunotherapy of hepatoma with a monoclonal antibody against murine endoglin

    Guang-Hong Tan; Feng-Ying Huang; Hua Wang; Yong-Hao Huang; Ying-Ying Lin; Yue-Nan Li

    2007-01-01

    AIM: To explore the capability of a monoclonal antibody(mAb) against murine endoglin to inhibit tumor angiogenesis and suppression of hepatoma growth in murine models.METHODS: A monoclonal antibody against murine endoglin was purified by affinity chromatography and passively transfused through tail veins in two murine hepatoma models. Tumor volume and survival time were observed at three-day intervals for 48 d. Microvessels in tumor tissues were detected by immunohistochemistry against CD31, and angiogenesis in vivo was determined by alginate encapsulated assay. In addition, tumor cell apoptosis was detected by TUNEL assay.RESULTS: Passive immunotherapy with anti-endoglin mAb could effectively suppress tumor growth, and prolonged the survival time of hepatoma-bearing mice.Angiogenesis was apparently inhibited within the tumor tissues, and the vascularization of alginate beads was also reduced in the mice passively transfused with antiendoglin mAb. In addition, increased apoptotic cells were observed within the tumor tissues from the mice passively transfused with anti-endoglin mAb.CONCLUSION: Passive immunotherapy with antiendoglin mAb effectively inhibits tumor growth via inhibiting tumor angiogenesis and increasing tumor cell apoptosis, which may be highly correlated with the blockage of endoglin-related signal pathway induced by anti-endoglin mAb.

  7. Murine myocardium OCT imaging with a blood substitute

    Kim, Jeehyun; Villard, Joseph W.; Lee, Ho; Feldman, Marc D.; Milner, Thomas E.

    2002-06-01

    Imaging of the in vivo murine myocardium using optical coherence tomography (OCT) is described. Application of conventional techniques (e.g. MRI, Ultrasound imaging) for imaging the murine myocardium is problematic because the wall thickness is less than 1.5mm (20g mouse), and the heart rate can be as high as six-hundred beats per minute. To acquire a real-time image of the murine myocardium, OCT can provide sufficient spatial resolution (10 micrometers ) and imaging speed (1000 A-Scans/s). Strong light scattering by blood in the heart causes significant light attenuation making delineation of the endocardium-chamber boundary problematic. By replacing whole blood in the mouse with an artificial blood substitute we demonstrate significant reduction of light scattering in the murine myocardium. The results indicate a significant reduction in light scattering as whole blood hematocrit is diminished below 5%. To measure thickness change of the myocardium during one cycle, a myocardium edge detection algorithm is developed and demonstrated.

  8. Pharmacodynamics of Doxycycline in a Murine Malaria Model▿

    Batty, Kevin T.; Law, Angela S. F.; Stirling, Verity; Moore, Brioni R.

    2007-01-01

    Doxycycline is reported to impair second-generation parasite schizogony. The effects of doxycycline alone and combined with dihydroartemisinin were investigated in a murine malaria model. Doxycycline lowered the rate of parasite growth within 2 days, with maximum effect in 6 days. Addition of dihydroartemisinin led to an additive antimalarial effect.

  9. Transplacental murine cytomegalovirus infection in the brain of SCID mice

    Jaquish Dawn V

    2007-03-01

    Full Text Available Abstract Background Congenital cytomegalovirus (CMV infection is the most common congenital viral infection in humans and the major nonhereditary cause of central nervous system (CNS developmental disorders. Previous attempts to develop a murine CMV (MCMV model of natural congenital human CMV (HCMV infection have failed because MCMV does not cross the placenta in immunocompetent mice. Results In marked contrast with immunocompetent mice, C.B-17 SCID (severe combined immunodeficient mice were found to be highly susceptible to natural MCMV transplacental transmission and congenital infection. Timed-pregnant SCID mice were intraperitoneally (IP injected with MCMV at embryonic (E stages E0-E7, and vertical MCMV transmission was evaluated using nested polymerase chain reaction (nPCR, in situ hybridization (ISH and immunohistochemical (IHC assays. SCID mouse dams IP injected at E0 with 102 PFU of MCMV died or resorbed their fetuses by E18. Viable fetuses collected at E18 from SCID mice IP injected with 102–104 PFU of MCMV at E7 did not demonstrate vertical MCMV transmission. Notably, transplacental MCMV transmission was confirmed in E18 fetuses from SCID mice IP injected with 103 PFU of MCMV at stages E3-E5. The maximum rate of transplacental MCMV transmission (53% at E18 occurred when SCID mouse dams were IP injected with 103 PFU of MCMV at E4. Congenital infection was confirmed by IHC immunostaining of MCMV antigens in 26% of the MCMV nPCR positive E18 fetuses. Transplacental MCMV transmission was associated with intrauterine growth retardation and microcephaly. Additionally, E18 fetuses with MCMV nPCR positive brains had cerebral interleukin-1α (IL-1α expression significantly upregulated and cerebral IL-1 receptor II (IL-1RII transcription significantly downregulated. However, MCMV-induced changes in cerebral cytokine expression were not associated with any histological signs of MCMV infection or inflammation in the brain. Conclusion Severe T

  10. Angiogenesis gene expression in murine endothelial cells during post-pneumonectomy lung growth

    Konerding Moritz A

    2011-07-01

    Full Text Available Abstract Although blood vessel growth occurs readily in the systemic bronchial circulation, angiogenesis in the pulmonary circulation is rare. Compensatory lung growth after pneumonectomy is an experimental model with presumed alveolar capillary angiogenesis. To investigate the genes participating in murine neoalveolarization, we studied the expression of angiogenesis genes in lung endothelial cells. After left pneumonectomy, the remaining right lung was examined on days 3, 6, 14 and 21days after surgery and compared to both no surgery and sham thoracotomy controls. The lungs were enzymatically digested and CD31+ endothelial cells were isolated using flow cytometry cell sorting. The transcriptional profile of the CD31+ endothelial cells was assessed using quantitative real-time polymerase chain reaction (PCR arrays. Focusing on 84 angiogenesis-associated genes, we identified 22 genes with greater than 4-fold regulation and significantly enhanced transcription (p

  11. Overexpression of mclca3 in airway epithelium of asthmatic murine models with airway inflammation

    ZHANG Hui-lan; HE Li

    2010-01-01

    Asthma is a worldwide prevalent disease that is a considerable health burden in many countries.1 In recent years, the airway epithelium is increasingly recognized as a central contributor to the pathogenesis of asthma.2 One of the most highly induced genes in epithelial cells in experimental allergic airway disease is the third murine calcium-activated chloride channel homologue (mclca3, alias gob-5). Its human homology protein is hCLCA1,3,4 which has been identified as clinically relevant molecules in diseases with secretory dysfunctions including asthma and cystic fibrosis. In initial studies, mclca3 was thought to be a member of calcium-activated chloride channel (CaCCs) family,whereas some new interesting reports suggest that the two mclca3 cleavage products cannot form an anion channel on their own but may instead act as extracellular signaling molecules with as yet unknown functions and interacting partners.5

  12. Polysaccharide-rich fraction of Agaricus brasiliensis enhances the candidacidal activity of murine macrophages

    Priscila Raquel Martins

    2008-05-01

    Full Text Available A polysaccharide-rich fraction (ATF of medicinal mushroom Agaricus brasiliensis was evaluated on the candidacidal activity, H2O2 and nitric oxide (NO production, and expression of mannose receptors by murine peritoneal macrophages. Mice received three intraperitoneal (i.p. injections of ATF and after 48 h their peritoneal resident macrophages were assayed against Candida albicans yeast forms. The treatment increased fungicidal activity and it was associated with higher levels of H2O2, whereas NO production was not affected. We also found that the treatment enhances mannose receptor expression by peritoneal macrophages, which are involved in the attachment and phagocytosis of non-opsonized microorganisms. Treatment of animals with ATF was able to enhance the clearance of C. albicans during the first 6 h after the experimental i.p. infection. Our results suggest that this extract can increase host resistance against some infectious agents through the stimulation of microbicidal activity of macrophages.

  13. Directed evolution and targeted mutagenesis to murinize Listeria monocytogenes internalin A for enhanced infectivity in the murine oral infection model.

    Monk, Ian R

    2010-01-01

    Internalin A (InlA) is a critical virulence factor which mediates the initiation of Listeria monocytogenes infection by the oral route in permissive hosts. The interaction of InlA with the host cell ligand E-cadherin efficiently stimulates L. monocytogenes entry into human enterocytes, but has only a limited interaction with murine cells.

  14. Single Amino Acid Insertion in Loop 4 Confers Amphotropic Murine Leukemia Virus Receptor Function upon Murine Pit1

    Lundorf, Mikkel D.; Pedersen, Finn Skou; O'Hara, Bryan; Pedersen, Lene

    1998-01-01

    Pit1 is the human receptor for gibbon ape leukemia virus (GALV) and feline leukemia virus subgroup B (FeLV-B), while the related human protein Pit2 is a receptor for amphotropic murine leukemia virus (A-MuLV). The A-MuLV-related isolate 10A1 can utilize both Pit1 and Pit2 as receptors. A stretch of...

  15. Mouse Models of Multiple Sclerosis: Experimental Autoimmune Encephalomyelitis and Theiler’s Virus-Induced Demyelinating Disease

    McCarthy, Derrick P.; Richards, Maureen H.; Miller, Stephen D.

    2012-01-01

    Experimental autoimmune encephalomyelitis (EAE) and Theiler’s Murine Encephalitis Virus-Induced Demyelinating Disease (TMEV-IDD) are two clinically relevant murine models of multiple sclerosis (MS). Like MS, both are characterized by mononuclear cell infiltration into the CNS and demyelination. EAE is induced by either the administration of myelin protein or peptide in adjuvant or by the adoptive transfer of encephalitogenic T cell blasts into naïve recipients. The relative merits of each of ...

  16. Role of the effector and regulatory arms of the adaptative immune response in the pathophysiology of experimental asthma

    Amor Carro, Óscar

    2014-01-01

    Classic murine models of experimental asthma based on intraperitoneal sensitization followed by airway challenge do not reflect the way in which humans acquire allergic disease to airborne allergens. The interaction of the airway mucosa with the allergens may be essential for the triggering of the subsequent immune response. In the present work, we developed a murine model of allergic disease based on primary airway exposure to antigen followed by continuous airway challenge. Foll...

  17. Safety and efficacy of factor IX gene transfer to skeletal muscle in murine and canine hemophilia B models by adeno-associated viral vector serotype 1

    Arruda, Valder R.; Schuettrumpf, Joerg; Herzog, Roland W; Nichols, Timothy C.; Robinson, Nancy; Lotfi, Yasmin; Mingozzi, Federico; Xiao, Weidong; Couto, Linda B.; High, Katherine A.

    2003-01-01

    Adeno-associated viral (AAV) vectors (serotype 2) efficiently transduce skeletal muscle, and have been used as gene delivery vehicles for hemophilia B and for muscular dystrophies in experimental animals and humans. Recent reports suggest that AAV vectors based on serotypes 1, 5, and 7 transduce murine skeletal muscle much more efficiently than AAV-2, with reported increases in expression ranging from 2-fold to 1000-fold. We sought to determine whether this increased efficacy could be observe...

  18. Evaluation of the antitumor activity of interleukin-12 in an experimental murine model of colorectal cancer induced by 1,2 dimethylhydrazine (DMH Estudio de la respuesta antitumoral de la interleucina-12 en cáncer de colon inducido mediante 1,2-dimetilhidracina (DMH

    S. Coca

    2005-09-01

    I (p=0.028 and group III (p = 0.019. Other parameters measured, such as biggest tumor size and total tumoral volume were found to be lower in group II, although no statistical differences were found between groups. Only 10% of rats in group I showed moderated/extensive NK cell infiltration, vs. 60% of rats in group II (p = 0.077 and 70% in group III (p = 0.02. Conclusion: The administration of DMH to rodents provides a reliable and consistent means of inducing CRC that may be suitable for the evaluation of anti-cancer therapies. Our findings suggest that IL-12 is effective against the development of experimental CRC. Its antineoplastic effect could be attributed to the stimulus of this cytokine on the intratumoral infiltration of NK cells.Objetivo: la interleucina (IL-12 es una citocina que estimula la proliferación y la actividad citotóxica de los linfocito T y las células natural killer (NK. En trabajos previos se ha observado una relación entre la infiltración intratumoral de células NK y una mayor supervivencia en carcinomas colorrectales (CCR. Existen evidencias de un efecto aditivo en la actividad inmunomoduladora de la asociación de IL-12 con IL-2. Así, nos hemos propuesto el estudio de la capacidad de respuesta inmune antitumoral, tras la administración sistémica de IL-12 sola o combinada con IL-2, en un modelo experimental de CCR inducidos mediante la administración de 1,2-dimetilhidracina (DMH. Método: sesenta y cinco ratas Wistar de 6 semanas a las que se administró en inyección subcutánea una dosis semanal de DMH a razón de 20 mg/kg de peso durante 26 semanas. Finalizado el periodo de inducción, los animales se distribuyeron aleatoriamente en tres grupos. I: grupo control. Grupo II, se administró IL-12 recombinante murina. Grupo III: se administró IL-12, combinada con IL-2. Las ratas se sacrificaron en la semana 30, estudiándose los siguientes parámetros: número y localización de tumores, tamaño y carga tumoral. Se realiz

  19. Broadband acoustic properties of a murine skull

    Estrada, Héctor; Rebling, Johannes; Turner, Jake; Razansky, Daniel

    2016-03-01

    It has been well recognized that the presence of a skull imposes harsh restrictions on the use of ultrasound and optoacoustic techniques in the study, treatment and modulation of the brain function. We propose a rigorous modeling and experimental methodology for estimating the insertion loss and the elastic constants of the skull over a wide range of frequencies and incidence angles. A point-source-like excitation of ultrawideband acoustic radiation was induced via the absorption of nanosecond duration laser pulses by a 20 μm diameter microsphere. The acoustic waves transmitted through the skull are recorded by a broadband, spherically focused ultrasound transducer. A coregistered pulse-echo ultrasound scan is subsequently performed to provide accurate skull geometry to be fed into an acoustic transmission model represented in an angular spectrum domain. The modeling predictions were validated by measurements taken from a glass cover-slip and ex vivo adult mouse skulls. The flexible semi-analytical formulation of the model allows for seamless extension to other transducer geometries and diverse experimental scenarios involving broadband acoustic transmission through locally flat solid structures. It is anticipated that accurate quantification and modeling of the skull transmission effects would ultimately allow for skull aberration correction in a broad variety of applications employing transcranial detection or transmission of high frequency ultrasound.

  20. Resveratrol-loaded nanocapsules inhibit murine melanoma tumor growth.

    Carletto, Bruna; Berton, Juliana; Ferreira, Tamara Nascimento; Dalmolin, Luciana Facco; Paludo, Katia Sabrina; Mainardes, Rubiana Mara; Farago, Paulo Vitor; Favero, Giovani Marino

    2016-08-01

    In this study, resveratrol-loaded nanocapsules were developed and its antitumor activity tested on a melanoma mice model. These nanocapsules were spherically-shaped and presented suitable size, negative charge and high encapsulation efficiency for their use as a modified-release system of resveratrol. Nanoencapsulation leads to the drug amorphization. Resveratrol-loaded nanoparticles reduced cell viability of murine melanoma cells. There was a decrease in tumor volume, an increase in the necrotic area and inflammatory infiltrate of melanoma when resveratrol-loaded nanocapsules were compared to free resveratrol in treated mice. Nanoencapsulation of resveratrol also prevented metastasis and pulmonary hemorrhage. This modified-release technology containing resveratrol can be used as a feasible approach in order to inhibit murine melanoma tumor growth. PMID:27070053

  1. Antibody responses against xenotropic murine leukemia virus-related virus envelope in a murine model.

    Natalia Makarova

    Full Text Available BACKGROUND: Xenotropic murine leukemia virus-related virus (XMRV was recently discovered to be the first human gammaretrovirus that is associated with chronic fatigue syndrome and prostate cancer (PC. Although a mechanism for XMRV carcinogenesis is yet to be established, this virus belongs to the family of gammaretroviruses well known for their ability to induce cancer in the infected hosts. Since its original identification XMRV has been detected in several independent investigations; however, at this time significant controversy remains regarding reports of XMRV detection/prevalence in other cohorts and cell type/tissue distribution. The potential risk of human infection, coupled with the lack of knowledge about the basic biology of XMRV, warrants further research, including investigation of adaptive immune responses. To study immunogenicity in vivo, we vaccinated mice with a combination of recombinant vectors expressing codon-optimized sequences of XMRV gag and env genes and virus-like particles (VLP that had the size and morphology of live infectious XMRV. RESULTS: Immunization elicited Env-specific binding and neutralizing antibodies (NAb against XMRV in mice. The peak titers for ELISA-binding antibodies and NAb were 1:1024 and 1:464, respectively; however, high ELISA-binding and NAb titers were not sustained and persisted for less than three weeks after immunizations. CONCLUSIONS: Vaccine-induced XMRV Env antibody titers were transiently high, but their duration was short. The relatively rapid diminution in antibody levels may in part explain the differing prevalences reported for XMRV in various prostate cancer and chronic fatigue syndrome cohorts. The low level of immunogenicity observed in the present study may be characteristic of a natural XMRV infection in humans.

  2. Cytomegalovirus infection of murine testicular interstitial Leydig cells.

    Baskar, J F; Stanat, S C; Huang, E S

    1983-01-01

    We studied the susceptibility of mouse testicular interstitial Leydig cells to cytomegalovirus both in vivo and in vitro. The in vivo studies included intratesticular and intraperitoneal infection of 6-week-old mice with murine cytomegalovirus (MCMV); the in vitro studies involved an MCMV-Leydig cell interaction using a Leydig tumor cell line (I-10). MCMV-specific antigens were detected in interstitial Leydig cells in sections of MCMV-inoculated testes by an indirect immunofluorescence test. ...

  3. Molecular and physiological alterations in murine ventricular dysfunction.

    Rockman, H A; Ono, S.; Ross, R.S.; Jones, L R; Karimi, M.; Bhargava, V.; Ross, J; Chien, K R

    1994-01-01

    The present study reports the development and characterization of a murine model of right ventricular dysfunction following graded constriction in the pulmonary artery via microsurgical approaches. To analyze in vivo ventricular function, a technique of x-ray contrast microangiography was developed to allow the quantitative analysis of ventricular volumes and of ejection fraction in normal and pressure-overloaded right ventricle. Severe, chronic pulmonary arterial banding for 14 days resulted...

  4. Pharmacodynamics of Fluconazole in a Murine Model of Systemic Candidiasis

    Louie, Arnold; Drusano, George L.; Banerjee, Partha; Liu, Qing-Feng; Liu, Weiguo; Kaw, Pamela; Shayegani, Mehdi; Taber, Harry; Miller, Michael H.

    1998-01-01

    In this study we defined the pharmacodynamic parameter that optimizes outcome in deep-seated Candida albicans infections treated with fluconazole. Using a murine model of systemic candidiasis, we conducted single-dose dose-ranging studies with fluconazole to determine the dosage of this drug that resulted in a 50% reduction in fungal densities (50% effective dose [ED50]) in kidneys versus the fungal densities in the kidneys of untreated controls. We found that the ED50 of fluconazole given in...

  5. Murine immunization by cesium-137 irradiation attenuated Schistosoma mansoni cercariae

    Cesium-137, becoming a more readily available ionizing gamma radiation source for laboratory use, was shown to effectively attenuate Schistosoma mansoni cercariae for vaccine production. In parallel comparison studies with the murine model, cesium-137 attenuated cercariae consistently afforded better protection than did the cobalt-60 prepared vaccine. Dose-response data indicated that the optimal total irradiation with cesium-137 was between 45 and 50 Krad

  6. Murine immunization by cesium-137 irradiation attenuated Schistosoma mansoni cercariae

    Stek, M. Jr.; Minard, P.; Cruess, D.F.

    1984-06-01

    Cesium-137, becoming a more readily available ionizing gamma radiation source for laboratory use, was shown to effectively attenuate Schistosoma mansoni cercariae for vaccine production. In parallel comparison studies with the murine model, cesium-137 attenuated cercariae consistently afforded better protection than did the cobalt-60 prepared vaccine. Dose-response data indicated that the optimal total irradiation with cesium-137 was between 45 and 50 Krad.

  7. Integration of murine leukemia virus DNA depends on mitosis.

    Roe, T.; Reynolds, T. C.; Yu, G.; Brown, P O

    1993-01-01

    In synchronized rat or mouse cells infected with Moloney murine leukemia virus (MLV), integration of viral DNA and production of viral proteins occur only after the cells traverse mitosis. Integration is blocked when cells are prevented from progressing through mitosis. Viral nucleoprotein complexes isolated from arrested cells contain full-length viral DNA and can integrate this viral DNA in vitro, showing that the block to integration in arrested cells is not due to a lack of mature integra...

  8. Specific receptor binding of staphylococcal enterotoxins by murine splenic lymphocytes.

    Buxser, S; Bonventre, P F; Archer, D L

    1981-01-01

    We describe a reliable assay to measure the specific binding of 125I-labeled staphylococcal enterotoxin A (SEA) by murine spleen cells. Toxin binding by lymphocytes was specific in that it was inhibited by unlabeled SEA but not by unrelated proteins. The biological activity of SEA (T-lymphocyte mitogenesis) correlated with toxin binding to splenic lymphocytes. In the presence of high concentrations of [125I]SEA, specific binding increased rapidly and approached saturation after 2 h. Toxin bin...

  9. Factors Influencing RBC Alloimmunization: Lessons Learned from Murine Models

    Ryder, Alex B.; Zimring, James C.; Hendrickson, Jeanne E

    2014-01-01

    Red blood cell (RBC) alloimmunization may occur following transfusion or pregnancy/delivery. Although observational human studies have described the immunogenicity of RBC antigens and the clinical significance of RBC alloantibodies, studies of factors influencing RBC alloimmunization in humans are inherently limited by the large number of independent variables involved. This manuscript reviews data generated in murine models that utilize transgenic donor mice, which express RBC-specific model...

  10. A Murine Model of Contact Lens–Associated Fusarium Keratitis

    Sun, Yan; Chandra, Jyotsna; Mukherjee, Pranab; Szczotka-Flynn, Loretta; Ghannoum, Mahmoud A.; Pearlman, Eric

    2010-01-01

    The 2006 outbreak of contact lens–associated Fusarium keratitis resulted in more than 300 cases in the United States in which a commercial lens care product was implicated. In the current study, Fusarium grown as biofilm on silicone hydrogel lenses induced keratitis in a murine model and severity of disease and survival of the organisms were dependent on MyD88, IL-1R1, and TLR4.

  11. Embryonic expression and cloning of the murine GATA-3 gene.

    K. M. George; Leonard, M W; Roth, M. E.; Lieuw, Ken; Kioussis, D; Grosveld, Frank; Engel, Douglas

    1994-01-01

    textabstractWe describe the embryonic expression pattern as well as the cloning and initial transcriptional regulatory analysis of the murine (m) GATA-3 gene. In situ hybridization shows that mGATA-3 mRNA accumulation is temporally and spatially regulated during early development: although found most abundantly in the placenta prior to 10 days of embryogenesis, mGATA-3 expression becomes restricted to specific cells within the embryonic central nervous system (in the mesencephalon, diencephal...

  12. Adjuvant and anti-inflammatory properties of cigarette smoke in murine allergic airway inflammation.

    Trimble, Nancy J; Botelho, Fernando M; Bauer, Carla M T; Fattouh, Ramzi; Stämpfli, Martin R

    2009-01-01

    The impact of cigarette smoke on allergic asthma remains controversial both clinically and experimentally. The objective of this study was to investigate, in a murine model, how cigarette smoke affects immune inflammatory processes elicited by a surrogate allergen. In our experimental design, mice were concurrently exposed to cigarette smoke and ovalbumin (OVA), an innocuous antigen that, unless introduced in the context of an adjuvant, induces inhalation tolerance. We show that cigarette smoke exposure has adjuvant properties, allowing for allergic mucosal sensitization to OVA. Specifically, concurrent exposure to cigarette smoke and OVA for 2 weeks led to airway eosinophilia and goblet cell hyperplasia. In vivo OVA recall challenge 1 month after the last smoke exposure showed that concurrent exposure to OVA and cigarette smoke induced antigen-specific memory. Robust eosinophilia and OVA-specific IgG1 and IgE characterized the ensuing inflammatory response. Mechanistically, allergic sensitization was, in part, granulocyte macrophage colony-stimulating factor (GM-CSF) dependent, as a significant reduction in BAL eosinophilia was observed in mice treated with an anti-GM-CSF antibody. Of note, continuous smoke exposure attenuated the OVA recall response; decreased airway eosinophilia was observed in mice continuously exposed to cigarette smoke compared with mice that ceased the smoke exposure protocol. In conclusion, we demonstrate experimentally that while cigarette smoke acts as an adjuvant allowing for allergic sensitization, it also attenuates the ensuing eosinophilic inflammatory response. PMID:18635815

  13. Differentiation of Murine Embryonic Stem Cells into Endothelial Cells

    F. Fathi

    2008-01-01

    Full Text Available Objective: In this investigation Murine Embryonic Stem (ES cells were differentiatedinto endothelial cells.Materials and Methods: Murine ES cells (CCE cell line exposed to Alpha-MEM medium containing 10% FBS for 4 days. Then obtained Flk-1 (Flk-1:Vascular Endothelial Growth Factor Receptor 2 Positive cells were cultuted inEndothelial Growth Medium-2 (EGM-2 until the last day of experiment. Differentiatedcells were evaluated by immunocytochemistry, RT-PCR and Tube FormationAssays.Results: When the ES cells cultured in collagen coated dishes containingAlpha-MEM & FBS, Flk-1 positive cells were obtained. After transfering Flk-1positive cells into fibronectin coated dishes containing EGM2, the cells wereassumed a relatively uniform endothelial cell morphology and could be propagatedand expanded. Immunocytochemical and RT-PCR analysis of differentiatedcells showed that they take up acetylated low-density lipoprotein (LDL, express Flk-1, CD31 and bind the BS-l lectin. When placed in Matrigel, these MurineES cell–derived endothelial cells formed capillary-like structures characteristicof endothelial cellsConclusion: ES cell–derived endothelial cells provide a novel means to examine the mechanisms of endothelial cell development, and may open up new therapeutic strategies.

  14. Cloning and characterization of a murine SIL gene

    Collazo-Garcia, N.; Scherer, P.; Aplan, P.D. [Roswell Park Cancer Institute, Buffalo, NY (United States)

    1995-12-10

    The human SIL gene is disrupted by a site-specific interstitial deletion in 25% of children with T-cell acute lymphoblastic leukemia. Since transcriptionally active genes are prone to recombination events, the recurrent nature of this lesion suggests that the SIL gene product is transcriptionally active in the cell type that undergoes this interstitial deletion and that the SIL gene product may play a role in normal lymphoid development. To facilitate studies of SIL gene function, we have cloned and characterized a murine SIL gene. The predicted murine SIL protein is 75% identical to the human gene, with good homology throughout the open reading frame. An in vitro translated SIL cDNA generated a protein slightly larger than the predicted 139-kDa protein. Although a prior report detected SIL mRNA expression exclusively in hematopoietic tissues, a sensitive RT-PCR assay demonstrated SIL expression to be ubiquitous, detectable in all tissues examined. Since the RT-PCR assay suggested that SIL mRNA expression was higher in rapidly proliferating tissues, we assayed SIL mRNA expression using a murine erythroleukemia model of terminal differentiation and found it to be dramatically decreased in conjunction with terminal differentiation. These studies demonstrate that the human SIL gene product is quite well conserved in rodents and suggest that the SIL gene product may play a role in cell proliferation. 26 refs., 6 figs.

  15. Optimal in vitro culture conditions for murine predominant immature CD8a+ dendritic cells

    NA Ning; XU Lin; CAO Kai-yuan; LUO Yun; YUAN Guang-qing; XIANG Peng; HONG Liang-qing; LI Shu-nong

    2009-01-01

    Background The prospects of using immature CD8a+ dendritic cells (DC2) to establish transplant immunologic tolerance and treatments for autoimmune diseases in the future are promising. However, the methods for inducing DC2 are still being explored. The present study was aimed to investigate the optimal in vitro conditions for preparing large numbers f predominant DC2 from murine bone marrow cells.Methods Three groups of bone marrow cells cultured under different conditions were examined, namely a cytokine-induced experimental group (cytokine group), a control group with a low concentration of granulocyte-macrophage colony stimulating factor (GM-CSF, low GM-CSF group) and a control group without ndogenous cytokines. The cytokine group was cultured with 5 ng/ml GM-CSF, 25 ng/ml Fit3 ligand (Flt3L), 20 ng/ml interleukin 4 (IL-4) and 100 ng/ml stem cell factor (SCF). The low GM-CSF control group was cultured with 0.4 ng/ml GM-CSF, 25 ng/ml FIt3L and 100 ng/ml SCF, without IL-4. The control group without exogenous cytokines was cultured without dditional cytokines. All cells were cultured at 37℃ under 5% CO2. On days 3, 7 and 16, 4-color flow cytometry was carried out to analyze the cell phenotypes, and the total cell numbers were counted to analyze the cell yields. Phase-contrast microscopy was used to observe the cell morphologies.Results The cytokine group exhibited higher proportions f typical immature CD8a+ DC, especially on day 3, but the total cell number and DC2 proportion decreased during prolonged culture. The low GM-CSF control group showed the same tendencies as the cytokine group on days 16 and 22, but produced higher total cell numbers (P <0.05) with lower DC2 proportions and cell numbers. The control group without exogenous cytokines spontaneously generated a certain proportion of DC2, but with low total cell and DC2 numbers that decreased rapidly, especially during prolonged culture (days 7 and 16, P <0.05).Conclusions Culture in the presence of 5 ng

  16. Murine eosinophil differentiation factor. An eosinophil-specific colony- stimulating factor with activity for human cells

    1986-01-01

    A purified murine lymphokine, eosinophil differentiation factor (EDF), was found to be a selective stimulus for the clonal proliferation and differentiation of murine eosinophil progenitor cells, establishing it as the murine eosinophil colony-stimulating factor (Eo-CSF). EDF was also active on human eosinophil progenitors and mature blood eosinophils, but had no effect on neutrophil or macrophage precursor cells, nor on blood neutrophils. In culture of human bone marrow cells, EDF stimulated...

  17. Optimization of Gene Transfection in Murine Myeloma Cell Lines using Different Transfection Reagents

    Shabani, Mahdi; Hemmati, Sheyda; Hadavi, Reza; Amirghofran, Zahra; Jeddi-Tehrani, Mahmood; Rabbani, Hodjatallah; Shokri, Fazel

    2010-01-01

    Purification and isolation of cellular target proteins for monoclonal antibody (MAb) production is a difficult and time-consuming process. Immunization of mice with murine cell lines stably transfected with genes coding for xenogenic target molecules is an alternative method for mouse immunization and MAb production. Here we present data on transfection efficiency of some commercial reagents used for transfection of murine myeloma cell lines. Little is known about transfectability of murine m...

  18. Inhibitory mechanism of peptides and antibodies targeting murine urokinase-type plasminogen activator

    Liu, Zhuo

    2012-01-01

    high affinity and specificity of mupain-1-16 makes it a suitable inhibitor for targeting of murine uPA in order to investigate the importance of uPA in murine disease models. Secondly, two high affinity monoclonal antibodies targeting murine uPA (mU1 and mU3) were studied. These antibodies showed...... be important for future tumour model studies and the development of more efficient inhibitors against uPA....

  19. Pathway analysis of Candida albicans survival and virulence determinants in a murine infection model.

    Becker, Jeffrey M; Kauffman, Sarah J; Hauser, Melinda; Huang, Liyin; Lin, Molly; Sillaots, Susan; Jiang, Bo; Xu, Deming; Roemer, Terry

    2010-12-21

    One potentially rich source of possible targets for antifungal therapy are those Candida albicans genes deemed essential for growth under the standard culture (i.e., in vitro) conditions; however, these genes are largely unexplored as drug targets because essential genes are not experimentally amenable to conventional gene deletion and virulence studies. Using tetracycline-regulatable promoter-based conditional mutants, we investigated a murine model of candidiasis in which repressing essential genes in the host was achieved. By adding doxycycline to the drinking water starting 3 days prior to (dox - 3D) or 2 days post (dox + 2D) infection, the phenotypic consequences of temporal gene inactivation were assessed by monitoring animal survival and fungal burden in prophylaxis and acute infection settings. Of 177 selected conditional shut-off strains tested, the virulence of 102 was blocked under both repressing conditions, suggesting that the corresponding genes are essential for growth and survival in a murine host across early and established infection periods. Among these genes were those previously identified as antifungal drug targets (i.e., FKS1, ERG1, and ERG11), verifying that this methodology can be used to validate potential new targets. We also identify genes either conditionally essential or dispensable for in vitro growth but required for survival and virulence, including those in late stage ergosterol synthesis, or early steps in fatty acid or riboflavin biosynthesis. This study evaluates the role of essential genes with respect to pathogen virulence in a large-scale, systems biology context, and provides a general method for gene target validation and for uncovering unexpected antimicrobial targets. PMID:21135205

  20. Effects of DMSO on Diminazene Efficacy in Experimental Murine T. brucei Infection

    K.I. Eghianruwa; Anika, S.M.

    2012-01-01

    This study evaluated the influence of dimethyl sulfoxide (DMSO) daily supplementation on diminazene treatment of trypanosomosis. Four groups of Trypanosoma brucei brucei infected rats received 7.0 mg/kg diminazene aceturate on day 7 post infection. Three of the four groups received different doses of DMSO (0.5, 1.0 and 2.0 g/kg, respectively) in addition to diminazene treatment. The changes in hematological parameters and the weights of liver, spleen and heart caused by T. brucei infection we...

  1. Combined immunotherapy of experimental murine T-lymphoma with antibody-targeted cytostatics and immunomodulating agents

    Jelínková, Miroslava; Šrogl, J.; Rossmann, P.; Novák, M.; Ulbrich, Karel; Strohalm, Jiří; Rozprimová, L.; Říhová, Blanka

    London: School of Pharmacy , University of London, 1995, s. 51. [International Symposium on Polymer Therapeutics: From Laboratory to Clinical Practice /1./. London (GB), 10.01.1996-12.01.1996] R&D Projects: GA AV ČR 720407; GA ČR GA307/93/0057

  2. Prevention of heterotopic ossification: an experimental study using a plasma expander in a murine model

    Zimmermann, Stefan M; Schwitter, Lukas W.; Scheyerer, Max J.; Jentzsch, Thorsten; Simmen, Hans-Peter; Werner, Clément M. L.

    2016-01-01

    Background Heterotopic ossification (HO) is a frequent complication following orthopedic and trauma surgery. It often leads to substantial morbidity as many affected patients suffer from pain and joint contractures. Current prophylactic measures include nonsteroidal anti-inflammatory drugs (NSAID) and local radiation. However, several disadvantages such as delayed fracture healing and impaired ossification have been reported. For this reason, a novel approach for prevention of HO was searched...

  3. Therapy of experimental murine brucellosis with streptomycin, co-trimoxazole, ciprofloxacin, ofloxacin, pefloxacin, doxycycline, and rifampin.

    Shasha, B; Lang, R.; Rubinstein, E

    1992-01-01

    Mice infected with Brucella melitensis were treated with streptomycin, co-trimoxazole, ciprofloxacin, doxycycline, and rifampin intraperitoneally and with ciprofloxacin, ofloxacin, pefloxacin, doxycycline, and rifampin orally for 14 to 21 days. Doxycycline- and rifampin-treated animals (either route) demonstrated a cure rate significantly better than that of controls. Longer therapy periods were associated with a significantly better outcome. Therapy failure was observed in all mice treated w...

  4. Therapeutic effect of hydroxychloroquine on colorectal carcinogenesis in experimental murine colitis.

    Yao, Junlin; Xie, Jiansheng; Xie, Binbin; Li, Yiran; Jiang, Liming; Sui, Xinbing; Zhou, Xiaoyun; Pan, Hongming; Han, Weidong

    2016-09-01

    Chronic inflammation in the intestine is a strong risk factor for colitis-associated colorectal cancer (CAC). Hydroxychloroquine (HCQ) is widely used as an anti-inflammatory drug in the treatment of immune-mediated inflammatory disorders and various tumors. However, little is known regarding the effects of HCQ on colitis-associated tumorigenesis. In this study, mice treated with HCQ showed a significant reduction in early-stage colitis following azoxymethane (AOM)/dextran sodium sulfate (DSS) administration, as well as a remarkable inhibition of colonic tumorigenesis and tumor growth at late stages of CAC. Mechanistically, the therapeutic effects of HCQ were attributed to inhibition of inflammatory responses and production of mutagenic reactive oxygen species (ROS) in immune cells and subsequent promotion of apoptosis and cell cycle arrest in tumor cells. Furthermore, we found that HCQ inhibited the production of inflammatory cytokines and ROS in response to toll-like receptor 4 (TLR4) activation in macrophages. Our data presented herein may help guide the clinical use of HCQ as a prevention and treatment strategy for CAC. PMID:27288548

  5. The murine gammaherpesvirus-68 chemokine-binding protein M3 inhibits experimental autoimmune encephalomyelitis

    Millward, Jason M; Holst, Peter J; Høgh-Petersen, Mette; Thomsen, Allan R; Christensen, Jan P; Owens, Trevor

    Chemokines are critical mediators of immune cell entry into the central nervous system (CNS), as occurs in neuroinflammatory disease such as multiple sclerosis. Chemokines are also implicated in the immune response to viral infections. Many viruses encode proteins that mimic or block chemokine ac...

  6. Local Th1/Th2 Cytokine Expression in Experimental Murine Vaginal Candidiasis

    Weixiang OUYANG; Shanjuan CHEN; Zhixiang LIU; Yan WU; Jiawen LI

    2008-01-01

    In order to investigate the expression of Th1 and Th2 cytokines in the vaginal candidiasis caused by Candida, the fungal vaginitis model was established in female ICR mice by intravaginal inoculation of suspension of C. albicans after the animals were pretreated with estradiol. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect the expression of IL-2, IL-4, IL-10 and TGF-β1 in the vagina in the mice of different groups at different time points after the beginning of the experiment. The average expression level of IL-2 mRNA in group D (estrogen-treated mice) was significantly higher than that in groups H (estrogen-untreated mice) and I (control group) on the day 2. The average expression level of IL-4 mRNA in group D was significantly higher than that in groups I and H on the day 5. The average expression level of IL-10 mRNA in group D was significantly higher than that in groups H and I from day 7 to 11. The average expression level of TGF-β1 mRNA in group D was significantly higher than that in groups H and I at all time points. It was concludes that the high-level expression of IL-2 mRNA during early infection was associated with clearance of mucosal C. albicans, and the high-level expression of IL-10 mRNA during late stage of the infection was related to susceptibility to infection. TGF-β1 may play a predominant role when the virtual absence of changes in other Th-type cytokines during infection.

  7. Lessons from probiotic-host interaction studies in murine models of experimental colitis

    Claes, Ingmar; De Keersmaecker, Sigrid; Vanderleyden, Jos; Lebeer, Sarah

    2011-01-01

    In inflammatory bowel diseases (IBD), it is known that besides genetic and environmental factors (e.g. diet, drugs, stress), the microbiota play an important role in the pathogenesis. Patients with IBD have an altered microbiota (dysbiosis) and therefore, probiotics, defined as 'live micro-organisms that when administered in adequate amounts can confer a health benefit on the host', have been suggested as nutritional supplements to restore these imbalances. The best response on probiotics amo...

  8. Validation of two reference genes for mRNA level studies of murine disease models in neurobiology

    Meldgaard, Michael; Fenger, Christina; Lambertsen, Kate Lykke; Pedersen, Mads Damgaard; Ladeby, Rune; Finsen, Bente

    2006-01-01

    Reverse transcription of extracted cellular RNA combined with real-time PCR is now an established method for sensitive detection and quantification of specific mRNA level changes in experimental models of neurological diseases. To neutralize the impact of experimental error and make quantification......, identification of an unaffected reference gene is necessary. In this report, we present our findings from evaluation and validation of the genes encoding hypoxanthine guanine phosphoribosyl transferase 1 (HPRT1) and glyceraldehyde phosphate dehydrogenase (GAPDH) as individual reference genes in mRNA level...... studies involving four murine neurological disease models. We find both genes are suitable as a reference gene with these four models, provided quantification of subtle changes are avoided. We furthermore demonstrate that above a certain threshold of test mRNA level changes and given high quality RNA...

  9. Microbiome and Asthma: What Have Experimental Models Already Taught Us?

    Bonamichi-Santos, R.; Aun, M. V.; Agondi, R. C.; Kalil, J; Giavina-Bianchi, P.

    2015-01-01

    Asthma is a chronic inflammatory disease that imposes a substantial burden on patients, their families, and the community. Although many aspects of the pathogenesis of classical allergic asthma are well known by the scientific community, other points are not yet understood. Experimental asthma models, particularly murine models, have been used for over 100 years in order to better understand the immunopathology of asthma. It has been shown that human microbiome is an important component in th...

  10. Infeção experimental por Trypanosoma brucei brucei em modelo murino e estudo da eficácia farmacológica do benznidazol

    Pereira, João Luís Gomes

    2013-01-01

    ABSTRACT - TRYPANOSOMA BRUCEI BRUCEI MURINE EXPERIMENTAL MURINE INFECTION AND STUDIES ON PHARMACOLOCICAL EFFECTIVENESS OF BENZNIDAZOLE - African Trypanosomiasis (AT) is a parasitic disease caused by several species of Trypanosoma, transmitted by diptera of the Glossina genus, also known as the tsetse flies. This disease affects humans and animals, in humans takes the name of Sleeping Sickness, and in animals takes the name of Nagana. Diagnosis can be performed by parasite visualization...

  11. A Murine Model of Candida glabrata Vaginitis Shows No Evidence of an Inflammatory Immunopathogenic Response.

    Nash, Evelyn E; Peters, Brian M; Lilly, Elizabeth A; Noverr, Mairi C; Fidel, Paul L

    2016-01-01

    Candida glabrata is the second most common organism isolated from women with vulvovaginal candidiasis (VVC), particularly in women with uncontrolled diabetes mellitus. However, mechanisms involved in the pathogenesis of C. glabrata-associated VVC are unknown and have not been studied at any depth in animal models. The objective of this study was to evaluate host responses to infection following efforts to optimize a murine model of C. glabrata VVC. For this, various designs were evaluated for consistent experimental vaginal colonization (i.e., type 1 and type 2 diabetic mice, exogenous estrogen, varying inocula, and co-infection with C. albicans). Upon model optimization, vaginal fungal burden and polymorphonuclear neutrophil (PMN) recruitment were assessed longitudinally over 21 days post-inoculation, together with vaginal concentrations of IL-1β, S100A8 alarmin, lactate dehydrogenase (LDH), and in vivo biofilm formation. Consistent and sustained vaginal colonization with C. glabrata was achieved in estrogenized streptozotocin-induced type 1 diabetic mice. Vaginal PMN infiltration was consistently low, with IL-1β, S100A8, and LDH concentrations similar to uninoculated mice. Biofilm formation was not detected in vivo, and co-infection with C. albicans did not induce synergistic immunopathogenic effects. This data suggests that experimental vaginal colonization of C. glabrata is not associated with an inflammatory immunopathogenic response or biofilm formation. PMID:26807975

  12. A Murine Model of Candida glabrata Vaginitis Shows No Evidence of an Inflammatory Immunopathogenic Response.

    Evelyn E Nash

    Full Text Available Candida glabrata is the second most common organism isolated from women with vulvovaginal candidiasis (VVC, particularly in women with uncontrolled diabetes mellitus. However, mechanisms involved in the pathogenesis of C. glabrata-associated VVC are unknown and have not been studied at any depth in animal models. The objective of this study was to evaluate host responses to infection following efforts to optimize a murine model of C. glabrata VVC. For this, various designs were evaluated for consistent experimental vaginal colonization (i.e., type 1 and type 2 diabetic mice, exogenous estrogen, varying inocula, and co-infection with C. albicans. Upon model optimization, vaginal fungal burden and polymorphonuclear neutrophil (PMN recruitment were assessed longitudinally over 21 days post-inoculation, together with vaginal concentrations of IL-1β, S100A8 alarmin, lactate dehydrogenase (LDH, and in vivo biofilm formation. Consistent and sustained vaginal colonization with C. glabrata was achieved in estrogenized streptozotocin-induced type 1 diabetic mice. Vaginal PMN infiltration was consistently low, with IL-1β, S100A8, and LDH concentrations similar to uninoculated mice. Biofilm formation was not detected in vivo, and co-infection with C. albicans did not induce synergistic immunopathogenic effects. This data suggests that experimental vaginal colonization of C. glabrata is not associated with an inflammatory immunopathogenic response or biofilm formation.

  13. Characterization of a Novel Murine Model to Study Zika Virus

    Rossi, Shannan L.; Tesh, Robert B.; Azar, Sasha R.; Muruato, Antonio E.; Hanley, Kathryn A.; Auguste, Albert J.; Langsjoen, Rose M.; Paessler, Slobodan; Vasilakis, Nikos; Weaver, Scott C.

    2016-01-01

    The mosquito-borne Zika virus (ZIKV) is responsible for an explosive ongoing outbreak of febrile illness across the Americas. ZIKV was previously thought to cause only a mild, flu-like illness, but during the current outbreak, an association with Guillain–Barré syndrome and microcephaly in neonates has been detected. A previous study showed that ZIKV requires murine adaptation to generate reproducible murine disease. In our study, a low-passage Cambodian isolate caused disease and mortality in mice lacking the interferon (IFN) alpha receptor (A129 mice) in an age-dependent manner, but not in similarly aged immunocompetent mice. In A129 mice, viremia peaked at ∼107 plaque-forming units/mL by day 2 postinfection (PI) and reached high titers in the spleen by day 1. ZIKV was detected in the brain on day 3 PI and caused signs of neurologic disease, including tremors, by day 6. Robust replication was also noted in the testis. In this model, all mice infected at the youngest age (3 weeks) succumbed to illness by day 7 PI. Older mice (11 weeks) showed signs of illness, viremia, and weight loss but recovered starting on day 8. In addition, AG129 mice, which lack both type I and II IFN responses, supported similar infection kinetics to A129 mice, but with exaggerated disease signs. This characterization of an Asian lineage ZIKV strain in a murine model, and one of the few studies reporting a model of Zika disease and demonstrating age-dependent morbidity and mortality, could provide a platform for testing the efficacy of antivirals and vaccines. PMID:27022155

  14. Nanomechanical phenotype of chondroadherin-null murine articular cartilage.

    Batista, Michael A; Nia, Hadi T; Önnerfjord, Patrik; Cox, Karen A; Ortiz, Christine; Grodzinsky, Alan J; Heinegård, Dick; Han, Lin

    2014-09-01

    Chondroadherin (CHAD), a class IV small leucine rich proteoglycan/protein (SLRP), was hypothesized to play important roles in regulating chondrocyte signaling and cartilage homeostasis. However, its roles in cartilage development and function are not well understood, and no major osteoarthritis-like phenotype was found in the murine model with CHAD genetically deleted (CHAD(-/-)). In this study, we used atomic force microscopy (AFM)-based nanoindentation to quantify the effects of CHAD deletion on changes in the biomechanical function of murine cartilage. In comparison to wild-type (WT) mice, CHAD-deletion resulted in a significant ≈70-80% reduction in the indentation modulus, Eind, of the superficial zone knee cartilage of 11 weeks, 4 months and 1 year old animals. This mechanical phenotype correlates well with observed increases in the heterogeneity collagen fibril diameters in the surface zone. The results suggest that CHAD mainly plays a major role in regulating the formation of the collagen fibrillar network during the early skeletal development. In contrast, CHAD-deletion had no appreciable effects on the indentation mechanics of middle/deep zone cartilage, likely due to the dominating role of aggrecan in the middle/deep zone. The presence of significant rate dependence of the indentation stiffness in both WT and CHAD(-/-) knee cartilage suggested the importance of both fluid flow induced poroelasticity and intrinsic viscoelasticity in murine cartilage biomechanical properties. Furthermore, the marked differences in the nanomechanical behavior of WT versus CHAD(-/-) cartilage contrasted sharply with the relative absence of overt differences in histological appearance. These observations highlight the sensitivity of nanomechanical tools in evaluating structural and mechanical phenotypes in transgenic mice. PMID:24892719

  15. Characterization of a Novel Murine Model to Study Zika Virus.

    Rossi, Shannan L; Tesh, Robert B; Azar, Sasha R; Muruato, Antonio E; Hanley, Kathryn A; Auguste, Albert J; Langsjoen, Rose M; Paessler, Slobodan; Vasilakis, Nikos; Weaver, Scott C

    2016-06-01

    The mosquito-borne Zika virus (ZIKV) is responsible for an explosive ongoing outbreak of febrile illness across the Americas. ZIKV was previously thought to cause only a mild, flu-like illness, but during the current outbreak, an association with Guillain-Barré syndrome and microcephaly in neonates has been detected. A previous study showed that ZIKV requires murine adaptation to generate reproducible murine disease. In our study, a low-passage Cambodian isolate caused disease and mortality in mice lacking the interferon (IFN) alpha receptor (A129 mice) in an age-dependent manner, but not in similarly aged immunocompetent mice. In A129 mice, viremia peaked at ∼10(7) plaque-forming units/mL by day 2 postinfection (PI) and reached high titers in the spleen by day 1. ZIKV was detected in the brain on day 3 PI and caused signs of neurologic disease, including tremors, by day 6. Robust replication was also noted in the testis. In this model, all mice infected at the youngest age (3 weeks) succumbed to illness by day 7 PI. Older mice (11 weeks) showed signs of illness, viremia, and weight loss but recovered starting on day 8. In addition, AG129 mice, which lack both type I and II IFN responses, supported similar infection kinetics to A129 mice, but with exaggerated disease signs. This characterization of an Asian lineage ZIKV strain in a murine model, and one of the few studies reporting a model of Zika disease and demonstrating age-dependent morbidity and mortality, could provide a platform for testing the efficacy of antivirals and vaccines. PMID:27022155

  16. IL-10 selectively regulates murine Ig isotype switching.

    Shparago, N; Zelazowski, P; Jin, L; McIntyre, T M; Stuber, E; Peçanha, L M; Kehry, M R; Mond, J J; Max, E E; Snapper, C M

    1996-05-01

    A role for IL-10 in regulating Ig isotype switching directly at the level of the murine B cell has not been previously reported. In this report we show that IL-10 selectively up-regulated IgM to IgG3 class switching in lipopolysaccharide (LPS)-activated cultures through a direct effect on membrane (m) IgM+IgG3(-)B cells in vitro. IL-10 stimulated a 3- to 4-fold enhancement (from 6-8 to 20-30%) in membrane mIgG3(+) cells and a significant increase in Smu-Sgamma3 DNA rearrangement events as measured by digestion-circularization PCR (DC-PCR) over that observed with LPS alone. IL-10 induction of switching to IgG3 was not accompanied by a corresponding increase in the steady-state levels of germline CHgamma3 RNA. By contrast, IL-10 strongly inhibited the transforming growth factor-beta-mediated generation of mIgA+ cells and Smu-Salpha DNA rearrangement events in LPS-, but not CD40 ligand (CD40L)-activated B cells. This effect was not accompanied by changes in the steady-state levels of germline CHalpha RNA. IL-10 had no effect on IL-4-mediated switching to either IgG1 or IgE in either LPS- or CD40L-activated B cells. Thus, IL-10 can either enhance or suppress switching to particular murine Ig isotypes but it differs from most other murine cytokines in that its effects on switching do not appear to be associated with changes in the corresponding steady-state levels of germline CH RNA. PMID:8671667

  17. Lipoxygenation of arachidonic acid by subcellular preparations from murine keratinocytes

    In these studies, we examined the possibility that cell-free preparations from murine keratinocytes possess 5-lipoxygenase activity in addition to the well-established cyclooxygenase pathway of arachidonic acid (AA) in these cells. Our data demonstrated that the high-speed (105,000 g) supernatant preparations of the murine keratinocytes metabolized [14C]AA into labeled lipoxygenase products. Portions of these radioactive metabolites cochromatographed and comigrated with 12-HETE (a marker for 12-lipoxygenase pathway) and with authentic LTB4 (a marker for 5-lipoxygenase pathway) on silicic acid column chromatography and by thin-layer chromatography (TLC) in two solvent systems respectively. Identity of the novel 14C which comigrated with LTB4 on both TLC and column chromatography was verified further by cochromatography of the free acid with authentic LTB4 on a reverse phase (RP) and the methyl esters on a straight phase high-pressure liquid chromatography. Incubation of the cell-free preparations with [14C]AA in the presence of ETYA, NDGA (inhibitors of cyclooxygenase and lipoxygenase pathways) as well as with 15-HETE (an inhibitor of lipoxygenase pathway) resulted in decreased formation of [14C] 12-HETE and the [14C]LTB4-like metabolite. On the contrary, incubations of the cell-free extracts with [14C] AA in the presence of indomethacin (a cyclooxygenase inhibitor) resulted in increased biosynthesis of the labeled lipoxygenase metabolites. These data indicate the existence of enzymes in soluble fraction of murine keratinocyte which can catalyze the transformation of [14C] AA into products of both the 12- and 5-lipoxygenase pathways

  18. Purification of Murine Monoclonal IgM Antibody

    1999-01-01

    This paper presents the purification of a monoclonal IgM antibody against human tumor associated antigen Lewis-Y by ion exchange chromatography and gel filtration.Enzyme-linked immunosorbent assay (ELISA) and SDS-polyacrylamide gel electrophoresis (PAGE) were used to identify purified IgM antibody.In flow cytometry analysis, the purified IgM antibody recognizes human breast tumor cell line MCF-7 which expresses Lewis-Y antigen.This work presents a new way for the purification of murine monoclonal IgM antibody.

  19. Dystrophic Spinal Deformities in a Neurofibromatosis Type 1 Murine Model

    Rhodes, Steven D.; Zhang, Wei; Yang, Dalong; Yang, Hao; Chen, Shi; Wu, Xiaohua; Li, Xiaohong; Yang, Xianlin; Mohammad, Khalid S.; Guise, Theresa A.; Amanda L Bergner; Stevenson, David A.; Yang, Feng-Chun

    2015-01-01

    Despite the high prevalence and significant morbidity of spinal anomalies in neurofibromatosis type 1 (NF1), the pathogenesis of these defects remains largely unknown. Here, we present two murine models: Nf1flox/−;PeriCre and Nf1flox/−;Col.2.3Cre mice, which recapitulate spinal deformities seen in the human disease. Dynamic histomorphometry and microtomographic studies show recalcitrant bone remodeling and distorted bone microarchitecture within the vertebral spine of Nf1flox/−;PeriCre and Nf...

  20. Genetic studies of the murine corneal response to Pseudomonas aeruginosa.

    Berk, R S; Beisel, K; Hazlett, L D

    1981-01-01

    The murine genetic control of resistance to Pseudomonas aeruginosa eye infection previously has been demonstrated to be regulated by two complementing dominant genes, PsCR1 and PsCR2. The PsCR1 locus apparently is not associated with the H-2 complex, whereas the PsCR2 locus could not definitively be associated with H-2. In this study we attempted to demonstrate a possible H-2 linkage of the PsCR2 locus. A panel of inbred congenic strains varying with either the H-2 haplotype or genetic backgr...

  1. DNA immunization confers protection against murine cytomegalovirus infection.

    González Armas, J C; Morello, C S; Cranmer, L D; Spector, D H

    1996-01-01

    The murine cytomegalovirus (MCMV) immediate-early gene 1 (IE1) encodes an 89-kDa phosphoprotein (pp89) which plays a key role in protecting BALB/c mice against the lethal effects of the MCMV infection. In this report, we have addressed the question of whether "naked DNA" vaccination with a eukaryotic expression vector (pcDNA-89) that contains the MCMV IE1 gene driven by a strong enhancer/promoter can confer protection. BALB/c mice were immunized intradermally with pcDNA-89 or with the plasmid...

  2. Production of antibodies which recognize opiate receptors on murine leukocytes

    Carr, D.J.J.; Bost, K.L.; Blalock, J.E.

    1988-01-01

    An antibody has been developed which recognizes opiate receptors on cells of the immune system. This antibody blocks specific binding of the radiolabeled opiate receptor ligand, /sup 3/H-dihydromorphine, to receptors on murine splenocytes. Additionally, the anti-receptor antibody competes with ..beta..-endorphin, meta-enkephalin, and naloxone for the same binding site on the leukocytes. Moreover, the anti-receptor antibody possesses agonist activity similar to ..beta..-endorphin in suppressing cAMP production by lymphocytes. These results suggest the development of an antibody which recognizes classical opiate receptors on cells of the immune system.

  3. DMPD: The actions of bacterial DNA on murine macrophages. [Dynamic Macrophage Pathway CSML Database

    Full Text Available 10534106 The actions of bacterial DNA on murine macrophages. Sester DP, Stacey KJ, ...Sweet MJ, Beasley SJ, Cronau SL, Hume DA. J Leukoc Biol. 1999 Oct;66(4):542-8. (.png) (.svg) (.html) (.csml) Show The... actions of bacterial DNA on murine macrophages. PubmedID 10534106 Title The actions of bacterial D

  4. Discovery of a Novel Murine Type C Retrovirus by Data Mining

    Bromham, Lindell; Clark, Francis; McKee, Jeff J.

    2001-01-01

    Analysis of genomic and expression data allows both identification and characterization of novel retroviruses. We describe a recombinant type C murine retrovirus, similar to the Mus dunni endogenous retrovirus, with VL30-like long terminal repeats and murine leukemia virus-like coding sequences. This virus is present in multiple copies in the mouse genome and expressed in a range of mouse tissues.

  5. Contribution of Candida albicans ALS1 to the Pathogenesis of Experimental Oropharyngeal Candidiasis

    Kamai, Yasuki; Kubota, Mikie; Kamai, Yoko; Hosokawa, Tsunemichi; Fukuoka, Takashi; Filler, Scott G.

    2002-01-01

    We investigated the contribution of Candida albicans ALS1, which encodes a candidal adhesin, to the pathogenesis of experimental murine oropharyngeal candidiasis. Our results indicate that the ALS1 gene product is important for the adherence of the organism to the oral mucosa during the early stage of the infection.

  6. Proteolytically modified human beta 2-microglobulin augments the specific cytotoxic activity in murine mixed lymphocyte culture

    Nissen, Mogens Holst; Claësson, M H

    1987-01-01

    (M-beta 2-m) bind to murine lymphocytes expressing H-2 class I antigens; M-beta 2-m, when added at day 0 and 1 of culture in nanomolar concentrations to a one-way murine allogeneic mixed lymphocyte culture (MLC) augments the generation of specific cytotoxic T lymphocytes; M-beta 2-m increases the...... endogenous production of interleukin 2 in the MLC culture; monoclonal antibody which reacts with both the native beta 2-m and M-beta 2-m molecule blocks the augmentation of cytotoxic T lymphocyte production induced by M-beta 2-m; murine as well as human MLC responder cells can proteolytically modify native...... human beta 2-m; and the modifying activity of murine MLC responder cells was blocked in an intermediary step by an alloantibody, which reacts specifically with murine major histocompatibility complex, class I-associated beta 2-m. These findings suggest that the modification process is preceded by an...

  7. The role of Hibiscus sabdariffa L. (Roselle) in maintenance of ex vivo murine bone marrow-derived hematopoietic stem cells.

    Abdul Hamid, Zariyantey; Lin Lin, Winnie Hii; Abdalla, Basma Jibril; Bee Yuen, Ong; Latif, Elda Surhaida; Mohamed, Jamaludin; Rajab, Nor Fadilah; Paik Wah, Chow; Wak Harto, Muhd Khairul Akmal; Budin, Siti Balkis

    2014-01-01

    Hematopoietic stem cells- (HSCs-) based therapy requires ex vivo expansion of HSCs prior to therapeutic use. However, ex vivo culture was reported to promote excessive production of reactive oxygen species (ROS), exposing HSCs to oxidative damage. Efforts to overcome this limitation include the use of antioxidants. In this study, the role of Hibiscus sabdariffa L. (Roselle) in maintenance of cultured murine bone marrow-derived HSCs was investigated. Aqueous extract of Roselle was added at varying concentrations (0-1000 ng/mL) for 24 hours to the freshly isolated murine bone marrow cells (BMCs) cultures. Effects of Roselle on cell viability, reactive oxygen species (ROS) production, glutathione (GSH) level, superoxide dismutase (SOD) activity, and DNA damage were investigated. Roselle enhanced the survival (P < 0.05) of BMCs at 500 and 1000 ng/mL, increased survival of Sca-1(+) cells (HSCs) at 500 ng/mL, and maintained HSCs phenotype as shown from nonremarkable changes of surface marker antigen (Sca-1) expression in all experimental groups. Roselle increased (P < 0.05) the GSH level and SOD activity but the level of reactive oxygen species (ROS) was unaffected. Moreover, Roselle showed significant cellular genoprotective potency against H2O2-induced DNA damage. Conclusively, Roselle shows novel property as potential supplement and genoprotectant against oxidative damage to cultured HSCs. PMID:25405216

  8. Multi-walled carbon nanotubes: biodegradation by gastric agents in vitro and effect on murine intestinal system

    Masyutin, A.; Erokhina, M.; Sychevskaya, K.; Gusev, A.; Vasyukova, I.; Smirnova, E.; Onishchenko, G.

    2015-11-01

    One of the main questions limiting application of fibrous carbon nanomaterials (CNM) in medicine and food industry concerns presumptive degradation of CNM in living organisms. In this study, we have investigated biodegradation of multi-walled carbon nanotubes (MWCNTs) by gastric agents in vitro and influence of ingested MWCNTs on murine intestine. Using scanning, conventional transmission and analytical electron microscopy, we demonstrated that industrial MWCNTs treated in vitro by 0.1 M hydrochloric acid (pH=1) and gastric juice (pH=2-3) isolated from murine stomach, are subjected to incomplete degradation. After 30 days of oral administration to experimental mice, we did find MWCNTs in the cells of small intestine, and it may indicate that agglomerates of MWCNTs do not penetrate into colon epithelia and do not accumulate in enterocytes. However, we observed local areas of necrotic damages of intestinal villi. It seems likely, therefore, that MWCNTs end up leaving gastrointestinal tract by excretion with the feces. Our results suggest that MWCNTs do not undergo complete degradation in gastrointestinal tract of mice, and passing through non-degraded particles may negatively affect intestinal system.

  9. Electrospun poly({epsilon}-caprolactone)/Ca-deficient hydroxyapatite nanohybrids: Microstructure, mechanical properties and cell response by murine embryonic stem cells

    Bianco, Alessandra, E-mail: bianco@stc.uniroma2.it [Department of Chemical Sciences and Technologies, Research Unit INSTM Tor Vergata, University of Rome Tor Vergata, via Ricerca Scientifica, 00133 Rome (Italy); Di Federico, Erica [Department of Chemical Sciences and Technologies, Research Unit INSTM Tor Vergata, University of Rome Tor Vergata, via Ricerca Scientifica, 00133 Rome (Italy); Moscatelli, Ilana; Camaioni, Antonella [Department of Public Health and Cell Biology, University of Rome Tor Vergata, via Montpellier, 00133 Rome (Italy); Armentano, Ilaria [Material Science and Technology Center, Research Unit INSTM, NIPLAB, University of Perugia, Terni (Italy); Campagnolo, Luisa [Department of Public Health and Cell Biology, University of Rome Tor Vergata, via Montpellier, 00133 Rome (Italy); Dottori, Mariaserena; Kenny, Jose Maria [Material Science and Technology Center, Research Unit INSTM, NIPLAB, University of Perugia, Terni (Italy); Siracusa, Gregorio [Department of Public Health and Cell Biology, University of Rome Tor Vergata, via Montpellier, 00133 Rome (Italy); Gusmano, Gualtiero [Department of Chemical Sciences and Technologies, Research Unit INSTM Tor Vergata, University of Rome Tor Vergata, via Ricerca Scientifica, 00133 Rome (Italy)

    2009-08-01

    Nanohybrid scaffolds mimicking extracellular matrix are promising experimental models to study stem cell behaviour, in terms of adhesion and proliferation. In the present study, the structural characterization of a novel electrospun nanohybrid and the analysis of cell response by a highly sensitive cell type, embryonic stem (ES) cells, are investigated. Ca-deficient hydroxyapatite nanocrystals (d-HAp) were synthesized by precipitation. Fibrous PCL/d-HAp nanohybrids were obtained by electrospinning, d-HAp content ranging between 2 and 55 wt.%. Electrospun mats showed a non-woven architecture, average fiber size was 1.5 {+-}0.5 {mu}m, porosity 80-90%, and specific surface area 16 m{sup 2} g{sup -1}. Up to 6.4 wt.% d-HAp content, the nanohybrids displayed comparable microstructural, mechanical and dynamo-mechanical properties. Murine ES cell response to neat PCL and to nanohybrid PCL/d-HAp (6.4 wt.%) mats was evaluated by analyzing morphological, metabolic and functional markers. Cells growing on either scaffold proliferated and maintained pluripotency markers at essentially the same rate as cells growing on standard tissue culture plates with no detectable signs of cytotoxicity, despite a lower cell adhesion at the beginning of culture. These results indicate that electrospun PCL scaffolds may provide adequate supports for murine ES cell proliferation in a pluripotent state, and that the presence of d-HAp within the mat does not interfere with their growth.

  10. Electrospun poly(ε-caprolactone)/Ca-deficient hydroxyapatite nanohybrids: Microstructure, mechanical properties and cell response by murine embryonic stem cells

    Nanohybrid scaffolds mimicking extracellular matrix are promising experimental models to study stem cell behaviour, in terms of adhesion and proliferation. In the present study, the structural characterization of a novel electrospun nanohybrid and the analysis of cell response by a highly sensitive cell type, embryonic stem (ES) cells, are investigated. Ca-deficient hydroxyapatite nanocrystals (d-HAp) were synthesized by precipitation. Fibrous PCL/d-HAp nanohybrids were obtained by electrospinning, d-HAp content ranging between 2 and 55 wt.%. Electrospun mats showed a non-woven architecture, average fiber size was 1.5 ±0.5 μm, porosity 80-90%, and specific surface area 16 m2 g-1. Up to 6.4 wt.% d-HAp content, the nanohybrids displayed comparable microstructural, mechanical and dynamo-mechanical properties. Murine ES cell response to neat PCL and to nanohybrid PCL/d-HAp (6.4 wt.%) mats was evaluated by analyzing morphological, metabolic and functional markers. Cells growing on either scaffold proliferated and maintained pluripotency markers at essentially the same rate as cells growing on standard tissue culture plates with no detectable signs of cytotoxicity, despite a lower cell adhesion at the beginning of culture. These results indicate that electrospun PCL scaffolds may provide adequate supports for murine ES cell proliferation in a pluripotent state, and that the presence of d-HAp within the mat does not interfere with their growth.

  11. Cloning and characterization of the murine claudin-5 promoter.

    Burek, Malgorzata; Förster, Carola Y

    2009-01-27

    Claudin-5, an integral tight junction protein component, plays a critical role in permeability of the endothelial cell barrier. Recently, we have shown that claudin-5 protein is down-regulated by the proinflammatory cytokine TNF alpha and its levels restored by dexamethasone treatment. In order to investigate the regulation of claudin-5 at the transcriptional level, we have cloned the murine claudin-5 promoter. The claudin-5 promoter sequence (1131 bp) showed no consensus TATA-box. We identified putative transcription factor binding sites, including six full and two half sites degenerated glucocorticoid-response elements (GREs), two NFkappaB, three Sp1, one Sp2, one Ap2, as well as three E-boxes. Serially deleted promoter constructs showed high basal activity. TNF alpha significantly reduced the promoter activity and mRNA levels of claudin-5 in brain cEND and myocardial MyEND endothelial cells. Dexamethasone treatment led to a significant increase of the murine claudin-5 promoter activity and mRNA levels in cEND cells. However, no claudin-5 induction could be observed in MyEND cells in response to dexamethasone. Our studies suggest tissue-specific regulation of the claudin-5 gene via glucocorticoids and a high vulnerability of claudin-5 to TNF alpha. This could be an important mechanism in diseases accompanied by the release of proinflammatory cytokines, for example in patients with chronic heart failure or multiple sclerosis. PMID:18996436

  12. Scanning electron microscopy of the neuropathology of murine cerebral malaria

    Brenneis Christian

    2006-11-01

    Full Text Available Abstract Background The mechanisms leading to death and functional impairments due to cerebral malaria (CM are yet not fully understood. Most of the knowledge about the pathomechanisms of CM originates from studies in animal models. Though extensive histopathological studies of the murine brain during CM are existing, alterations have not been visualized by scanning electron microscopy (SEM so far. The present study investigates the neuropathological features of murine CM by applying SEM. Methods C57BL/6J mice were infected with Plasmodium berghei ANKA blood stages. When typical symptoms of CM developed perfused brains were processed for SEM or light microscopy, respectively. Results Ultrastructural hallmarks were disruption of vessel walls, parenchymal haemorrhage, leukocyte sequestration to the endothelium, and diapedesis of macrophages and lymphocytes into the Virchow-Robin space. Villous appearance of observed lymphocytes were indicative of activated state. Cerebral oedema was evidenced by enlargement of perivascular spaces. Conclusion The results of the present study corroborate the current understanding of CM pathophysiology, further support the prominent role of the local immune system in the neuropathology of CM and might expose new perspectives for further interventional studies.

  13. Immunological impact of magnetic nanoparticles (Ferucarbotran) on murine peritoneal macrophages

    Yeh, Chen-Hao [National Taiwan University, Department of Horticulture (China); Hsiao, Jong-Kai [National Taiwan University Hospital and College of Medicine, Department of Medical Imaging (China); Wang, Jaw-Lin [National Taiwan University, Institute of Biomedical Engineering (China); Sheu, Fuu, E-mail: fsheu@ntu.edu.t [National Taiwan University, Department of Horticulture (China)

    2010-01-15

    Ferucarbotran, a clinically used superparamagnetic iron oxide, is widely developed as a magnetic resonance imaging (MRI) contrast agent and has the potential to improve the monitoring of macrophage recirculation in vivo. However, the biological effect of Ferucarbotran or magnetic nanoparticles (MNPs) on macrophage is not clearly understood yet. This study is aimed to examine the immunological impact of Ferucarbotran toward murine peritoneal macrophages. Cells treated with Ferucarbotran demonstrated a dose-responsive increase of granularity in the cytoplasm. After 24 h of incubation, viability and cytotoxicity in macrophages treated with 200 {mu}g Fe/mL of Ferucarbotran were not affected. Macrophages loaded with Ferucarbotran above 100 {mu}g Fe/mL showed a significant (p < 0.01) increase in cytokine (TNF-{alpha}, IL-1{beta}, IL-6) secretion and mRNA expression, followed by nitric oxide (NO) secretion and iNOS mRNA expression. Chemotactic responses of Ferucarbotran-preloaded macrophages toward CX3CL1 were significantly (p < 0.05) lower than those of untreated macrophages. Taking together, Ferucarbotran at high dose (100 {mu}g Fe/mL) could induce murine peritoneal macrophages activation in pro-inflammatory cytokine secretion and NO production.

  14. Great efficacy of sulfachloropyrazine-sodium against acute murine toxoplasmosis

    Yan-Bo Zeng; Bing Huang; Shun-Hai Zhu; Hui Dong; Hong-Yu Han; Lian-Lian Jiang; Quan Wang; Jun Cheng; Qi-Ping Zhao; Wei-Jiao Ma

    2012-01-01

    Objective: To identify more effective and less toxic drugs to treat animal toxoplasmosis.Methods:Efficacy of seven kinds of sulfonamides against Toxoplasma gondii (T. gondii) in an acute murine model was evaluated. The mice used throughout the study were randomly assigned to many groups (10 mice each), which either remained uninfected or were infected intraperitoneally with tachyzoites of T. gondii (strains RH and CN). All groups were then treated with different sulfonamides and the optimal treatment protocol was determined candidates. Sulfadiazine-sodium (SD) was used for comparison. Results: The optimal therapy involved gavaging mice twice per day with 250 mg/kg bw of sulfachloropyrazine-sodium (SPZ) for five days. Using this protocol, the average survival time and the time-point of 50% fatalities were prolonged significantly compared with SD treatment. Treatment with SPZ protected 40% of mice from death, and the heart and kidney tissue of these animals was parasite-free, as determined by nested-PCR. SPZ showed excellent therapeutic effects in the treatment of T. gondii in an acute murine model and is therefore a promising drug candidate for the treatment and prevention of T. gondii in animals. Conclusions: It can be concluded that the effective drug sulfachloropyrazine may be the new therapeutic options against animal toxoplasmosis.

  15. Differential chemokine responses in the murine brain following lyssavirus infection.

    Hicks, D J; Núñez, A; Banyard, A C; Williams, A; Ortiz-Pelaez, A; Fooks, A R; Johnson, N

    2013-11-01

    The hallmark of lyssavirus infection is lethal encephalomyelitis. Previous studies have reported distinct lyssavirus isolate-related differences in severity of cellular recruitment into the encephalon in a murine model of infection following peripheral inoculation with rabies virus (RABV) and European bat lyssavirus (EBLV)-1 and -2. In order to understand the role of chemokines in this process, comparative studies of the chemokine pattern, distribution and production in response to infection with these lyssaviruses were undertaken. Expression of CCL2, CCL5 and CXCL10 was observed throughout the murine brain with a distinct caudal bias in distribution, similar to both inflammatory changes and virus antigen distribution. CCL2 immunolabelling was localized to neuronal and astroglial populations. CCL5 immunolabelling was only detected in the astroglia, while CXCL10 labelling, although present in the astroglia, was more prominent in neurons. Isolate-dependent differences in the amount of chemokine immunolabelling in specific brain regions and chemokine production by neurons in vitro were observed, with a greater expression of CCL5 in vivo and CXCL10 production in vitro after EBLV infection. Additionally, strong positive associations between chemokine immunolabelling and perivascular cuffing and, to a lesser extent, virus antigen score were also observed. These differences in chemokine expression may explain the variation in severity of encephalitic changes observed in animals infected with different lyssavirus isolates. PMID:23746482

  16. Bifidobacteria DNA Induces Murine Macrophages Activation in vitro

    Yalin Li; Xun Qu; Hua Yang; Li Kang; Yingping Xu; Bo Bai; Wengang Song

    2005-01-01

    Previous studies have shown that oligodeoxynucleotides containing unmethylated CpG motifs were used as adjuvants for immunoregulation and immune response. This study was to explore the activation effects of Bifidobacteria DNA containing unmethylated CpG motifs (CpG DNA) on murine macrophage J774A.1 cells. The genomic DNA of Bifidobacteria was extracted and purified, and the methylation degree of CpG motifs was tested.The phagocytic ability of the macrophages was detected by flow cytometry. The cytokines (IL-1β, IL-6, IL-12p40 and TNF-α) levels in the culture supernatants of Bifidobacteria DNA treated J774A.1 cells were assayed by ELISA. The content of nitric oxide (NO) was detected by Griess reagent. After treated with Bifidobacteria DNA for 24h,Nile Red stain increased in J774A.1 macrophage, which suggested that the lipid metabolism increased in the macrophages. The phagocytic ability and levels of NO and cytokines of IL-1β, IL-6, IL-12p40 and TNF-α were significantly higher than PBS group and CT DNA group. The results indicated that Bifidobacteria DNA could activate murine macrophages J774A.1, which could provide scientific basis for the research and application of microorganism DNA preparation.

  17. Effect of blockage of costimulatory signal on murine abortion-prone model

    ZHAO Fu-xi; ZHANG Yuan-yuan; LIU Run-hua; LI Shuan-ming

    2007-01-01

    Background Inhibition of the key costimulatory signals results in T cell anergy, indicating the alloantigen-specific immunologic unresponsiveness. In this study, the effect of blockage of costimulatory signal CD86 on murine abortion-prone model was studied.Methods Thirty CBA/J female mice cohabitated with DBA/2 male or BALB/c male mice were investigated. CBA/J ×DBA/2 matings were used as the abortion-prone model, and CBA/J × BALB/c matings were used as the normal pregnant model. The abortion-prone models were divided into experimental and control groups, and the normal pregnant models were set as a normal group (10 mice in each group). The mice in the experimental group were treated with anti-mouse CD86 monoclonal antibody (mAb) (100 μg) on day 4.5 of gestation, while the controls received irrelevant-isotype matched rat IgG2b. As for the normal group, nothing was given to the mice. The mice were killed on day 13.5 of gestation, embryo resorption rate and the expression of transforming growth factor β1 (TGF-β1), plasminogen activator inhibitor 1 (PAI-1), and matrix metalloproteinase 9 (MMP-9) were detected. Then the data were analyzed by Chi-square test and Fisher's exact test.Results The embryo resorption rate in the experimental (8.2%) and normal groups (7.7%) was significantly lower than that of the control (23.5%, P<0.05). No significant difference was detected between the experimental and normal groups (P>0.05). The positive expression rates of TGF-β1 and PAI-1 proteins in the experimental and normal groups were significantly higher than those in the control group (P<0.05). The positive expression rate of MMP-9 protein in the experimental and normal groups was significantly lower than that in the control group (P<0.05). No significant difference in the positive expression rates of the three proteins was detected between the experimental and normal groups (P>0.05).Conclusions Blockage of costimulatory signal CD86 at early pregnancy can treat

  18. Expression of murine Fc receptors for IgG.

    Schreiber, R E; Buku, A; Unkeless, J C

    1990-06-15

    There are two distinct genes that encode murine low affinity Fc gamma RII, murine Fc gamma RII alpha, and murine Fc gamma RII beta, which are transcribed in specific cell lineages. Fc gamma RII alpha transcripts are present in macrophages, NK cells, and mesangial cells; Fc gamma RII beta transcripts are expressed in Fc gamma R-bearing B cells, T cells, and macrophages. We have devised a sandwich ELISA to quantify the expression of Fc gamma RII alpha protein. The ELISA is specific for Fc gamma RII alpha, and does not detect the closely related Fc gamma RII beta protein. Upon stimulation with IFN-gamma the Fc gamma RII beta- macrophage cell line J774a expressed a twelvefold enhanced level of Fc gamma RII alpha protein. Peritoneal macrophages synthesized varying amounts of Fc gamma RII alpha. High levels of Fc gamma RII alpha were observed in resident and thioglycollate-elicited peritoneal macrophages, but no Fc gamma RII alpha was detected in Bacillus Calmette Guérin-elicited macrophages. J774a cells stimulated with rIL-6 bound approximately twice as much anti-Fc gamma RII mAb 2.4G2 IgG as did unstimulated controls. However, the Fc gamma RII alpha-specific ELISA showed no change in the amount of Fc gamma RII alpha expressed. A probe encompassing the extracellular coding sequence of Fc gamma RII beta hybridized to two distinct transcripts that were elevated in rIL-6-stimulated J774a cells. One of these transcripts had the same mobility in electrophoresis as Fc gamma RII alpha mRNA and hybridized to an Fc gamma RII alpha-specific probe, whereas the other transcript was larger and did not hybridize to probes specific for either Fc gamma RII alpha or Fc gamma RII beta. Moreover, we confirmed, with an Fc gamma RII beta-specific probe, that J774a cells do not make Fc gamma RII beta mRNA. Thus, the larger transcript appears to encode a novel Fc gamma RII. We suggest that the increased level of binding of the anti-Fc gamma RII mAb 2.4G2 to rIL-6-induced cells represents

  19. Experimental philosophy.

    Knobe, Joshua; Buckwalter, Wesley; Nichols, Shaun; Robbins, Philip; Sarkissian, Hagop; Sommers, Tamler

    2012-01-01

    Experimental philosophy is a new interdisciplinary field that uses methods normally associated with psychology to investigate questions normally associated with philosophy. The present review focuses on research in experimental philosophy on four central questions. First, why is it that people's moral judgments appear to influence their intuitions about seemingly nonmoral questions? Second, do people think that moral questions have objective answers, or do they see morality as fundamentally relative? Third, do people believe in free will, and do they see free will as compatible with determinism? Fourth, how do people determine whether an entity is conscious? PMID:21801019

  20. Effect of Cold Plasma on Cell Viability and Collagen Synthesis in Cultured Murine Fibroblasts

    Shi, Xingmin; Cai, Jingfen; Xu, Guimin; Ren, Hongbin; Chen, Sile; Chang, Zhengshi; Liu, Jinren; Huang, Chongya; Zhang, Guanjun; Wu, Xili

    2016-04-01

    An argon atmospheric pressure plasma jet was employed to treat L929 murine fibroblasts cultured in vitro. Experimental results showed that, compared with the control cells, the treatment of fibroblasts with 15 s of plasma led to a significant increase of cell viability and collagen synthesis, while the treatment of 25 s plasma resulted in a remarkable decrease. Exploration of related mechanisms suggested that cold plasma could up-regulate CyclinD1 gene expression and down-regulate p27 gene expression at a low dose, while it could down-regulate CyclinD1 expression and up-regulate p27 expression at a higher dose, thus altering the cell cycle progression, and then affecting cell viability and collagen synthesis of fibroblasts. supported partly by National Natural Science Foundation of China (Nos. 81372076, 51307133 and 51221005), China National Funds for Distinguished Young Scientists (No. 51125029), the Sci-Tech Project of Shaanxi Province of China (No. 2010K16-04), and the Fundamental Research Funds for the Central Universities of China (No. xkjc2013004)

  1. Murine scrapie infection causes an abnormal germinal centre reaction in the spleen.

    McGovern, G; Brown, K L; Bruce, M E; Jeffrey, M

    2004-01-01

    Follicular dendritic cells (FDCs) of the lymphoreticular system play a role in the peripheral replication of prion proteins in some transmissible spongiform encephalopathies (TSEs), including experimental murine scrapie models. Disease-specific PrP (PrPd) accumulation occurs in association with the plasmalemma and extracellular space around FDC dendrites, but no specific immunological response has yet been reported in animals affected by TSEs. In the present study, morphology (light microscopical and ultrastructural) of secondary lymphoid follicles of the spleen were examined in mice infected with the ME7 strain of scrapie and in uninfected control mice, with or without immunological stimulation with sheep red blood cells (SRBCs), at 70 days post-inoculation or at the terminal stage of disease (268 days). Scrapie infection was associated with hypertrophy of FDC dendrites, increased retention of electron-dense material at the FDC plasma membrane, and increased maturation and numbers of B lymphocytes within secondary follicles. FDC hypertrophy was particularly conspicuous in immune-stimulated ME7-infected mice. The electron-dense material was associated with PrP Napoli accumulation, as determined by immunogold labelling. We hypothesize that immune system changes are associated with increased immune complex trapping by hypertrophic FDCs expressing PrP Napoli molecules at the plasmalemma of dendrites, and that this process is exaggerated by immune system stimulation. Contrary to previous dogma, these results show that a pathological response within the immune system follows scrapie infection. PMID:15003476

  2. Prolactin, systemic lupus erythematosus, and autoreactive B cells: lessons learnt from murine models.

    Saha, Subhrajit; Tieng, Arlene; Pepeljugoski, K Peter; Zandamn-Goddard, Gisele; Peeva, Elena

    2011-02-01

    The predominant prevalence of autoimmune diseases in women of reproductive age has led to the investigation of the effects of sex hormones on immune regulation and in autoimmune diseases, in particular the prototypic systemic autoimmune disease lupus. The female hormone prolactin has receptors beyond the reproductive axis including immune cells, and it is thought to promote autoimmunity in human and murine lupus. Induced hyperprolactinemia in experimental lupus models, regardless of gender, exacerbates disease activity and leads to premature death. Prolactin treatment in mice that are not prone to develop lupus leads to the development of a lupus-like phenotype. Persistent mild-moderate hyperprolactinemia alters the selection of the naïve B cell repertoire. Recent studies demonstrate that prolactin impairs all three mechanisms of B cell tolerance induction (negative selection, receptor editing, and anergy) and thereby contributes to the pathogenesis of autoimmunity. The effects of prolactin are genetically determined as shown by the differential response to the hormone in the different mice strains. Bromocriptine, a drug that inhibits prolactin secretion, abrogates some of the immune effects of this hormone. Further research is required to elucidate molecular mechanisms involved in immune effects of prolactin and to develop novel targeted treatments for SLE patients with prolactin-responsive disease. PMID:19937157

  3. Toll-Like Receptor-4 Dependent Small Intestinal Immune Responses Following Murine Arcobacter Butzleri Infection.

    Heimesaat, Markus M; Karadas, Gül; Fischer, André; Göbel, Ulf B; Alter, Thomas; Bereswill, Stefan; Gölz, Greta

    2015-12-01

    Sporadic cases of gastroenteritis have been attributed to Arcobacter butzleri infection, but information about the underlying immunopathological mechanisms is scarce. We have recently shown that experimental A. butzleri infection induces intestinal, extraintestinal and systemic immune responses in gnotobiotic IL-10(-/-) mice. The aim of the present study was to investigate the immunopathological role of Toll-like Receptor-4, the receptor for lipopolysaccharide and lipooligosaccharide of Gram-negative bacteria, during murine A. butzleri infection. To address this, gnotobiotic IL-10(-/-) mice lacking TLR-4 were generated by broad-spectrum antibiotic treatment and perorally infected with two different A. butzleri strains isolated from a patient (CCUG 30485) or fresh chicken meat (C1), respectively. Bacteria of either strain stably colonized the ilea of mice irrespective of their genotype at days 6 and 16 postinfection. As compared to IL-10(-/-) control animals, TLR-4(-/-) IL-10(-/-) mice were protected from A. butzleri-induced ileal apoptosis, from ileal influx of adaptive immune cells including T lymphocytes, regulatory T-cells and B lymphocytes, and from increased ileal IFN-γ secretion. Given that TLR-4-signaling is essential for A. butzleri-induced intestinal inflammation, we conclude that bacterial lipooligosaccharide or lipopolysaccharide compounds aggravate intestinal inflammation and may thus represent major virulence factors of Arcobacter. Future studies need to further unravel the molecular mechanisms of TLR-4-mediated A. butzleri-host interactions. PMID:26716022

  4. In vivo tumor localization using tumor-specific monkey xenoantibody, alloantibody, and murine monoclonal xenoantibody

    Specific in vivo localization of antibodies reactive with human melanoma cell membrane tumor associated antigens (TAA) has been attempted using congenitally athymic nude mice bearing subcutaneous human melanoma tumor xenografts as the experimental model. IgG fractions were prepared from each of several immune and control sera. Antimelanoma antibody sources included human alloantibody obtained from melanoma patients immunized against allogeneic melanoma cells, a monkey antiserum raised by immunization against a single human melanoma cell line, and a murine monoclonal antimelanoma antibody-secreting hybridoma cell line. Localization of these radiolabeled antibodies and of control IgG preparations to tumor tissue was determined by whole body scintigraphy and by differential tissue counting. Compared with the different control IgG preparations, each of the antimelanoma IgG preparations exhibited significant specific accumulation within the melanoma tissue. However, variation existed in the ability of each antimelanoma IgG to tumor preparation to localize despite attempts to control model parameters such as tumor source, in vivo passage number and mass. This variation appears to reflect basic biologic differences between tumors in different animals and possibly differences in the antigen-binding capacities of each IgG preparation following radioiodination. This technique for tumor localization is very promising and has obvious potential for clinical application

  5. Thrombin detection in murine plasma using engineered fluorescence resonance energy transfer aptadimers

    Trapaidze, Ana; Brut, Marie; Mazères, Serge; Estève, Daniel; Gué, Anne-Marie; Bancaud, Aurélien

    2015-12-01

    Biodetection strategies, in which two sides of one target protein are targeted simultaneously, have been shown to increase specificity, selectivity, and affinity, and it has been suggested that they constitute excellent candidates for protein sensing in complex media. In this study we propose a method to engineer the sequence of a DNA construct dedicated to reversible thrombin detection. This construct, called Fluorescence Resonance Energy Transfer (FRET) aptadimer, is assembled with two aptamers, which target different epitopes of thrombin, interconnected with a DNA linker that contains a FRET couple and a reversible double helix stem. In the absence of target, the stem is stable maintaining a FRET couple in close proximity, and fluorescence is unquenched upon thrombin addition due to the dehybridization of the stem. We define design rules for the conception of FRET aptadimers, and develop a software to optimize their functionality. One engineered FRET aptadimer sequence is subsequently characterized experimentally by temperature scanning fluorimetry, demonstrating the relevance of our technology for thrombin sensing in bulk and diluted murine plasma.

  6. 125I interstitial implants in the RIF-1 murine flank tumor: an animal model for brachytherapy

    The development of a model for interstitial brachytherapy that uses high-activity, removable 125I sources in the RIF-1 murine flank tumor is reported. Experimental end points are clonogenic cell and tumor regrowth delay assays. For the clonogenic cell assay, interestitial radiation is delivered at total doses of 500-10,000 rad at dose rates of 0.9-2.7 rad/min to cells in annuli of tissue in the tumor. Dose-survival curves are characterized by an initial shoulder followed by a straight (exponential) portion, with D0 similar to that of the curve obtained by external irradiation of the RIF-1 tumor in a self-contained cesium irradiator at similar dose rates. Tumor regrowth curves have been obtained for minimum tumor doses of 500-5000 rad; marked tumor regression has been observed with minimum tumor doses as low as 2000 rad, but results are not as reproducible as the results obtained with the clonogenic cell assay

  7. The Rho kinase inhibitor Fasudil up-regulates astrocytic glutamate transport subsequent to actin remodelling in murine cultured astrocytes

    Lau, Cl; O'Shea, Rd; Bischof, L;

    2011-01-01

    BACKGROUND AND PURPOSE Glutamate transporters play a major role in maintaining brain homeostasis and the astrocytic transporters, EAAT1 and EAAT2, are functionally dominant. Astrocytic excitatory amino acid transporters (EAATs) play important roles in various neuropathologies wherein astrocytes...... undergo cytoskeletal changes. Astrocytic plasticity is well documented, but the interface between EAAT function, actin and the astrocytic cytoskeleton is poorly understood. Because Rho kinase (ROCK) is a key determinant of actin polymerization, we investigated the effects of ROCK inhibitors on EAAT...... activity and astrocytic morphology. EXPERIMENTAL APPROACH The functional activity of glutamate transport was determined in murine cultured astrocytes after exposure to the ROCK inhibitors Fasudil (HA-1077) and Y27632 using biochemical, molecular and morphological approaches. Cytochemical analyses assessed...

  8. Monoclonal antibody to murine PECAM-1 (CD31) blocks acute inflammation in vivo

    1994-01-01

    A murine model of peritonitis was used to test the role of platelet/endothelial cell adhesion molecule 1 (PECAM-1/CD31) in acute inflammation. A monoclonal antibody (mAb) specific for murine PECAM-1 injected intravenously 4 h before the intraperitoneal injection of thioglycollate broth blocked leukocyte emigration into the peritoneal cavity for up to 48 h. This block was particularly evident for neutrophils. Control mAb, including one that bound to murine CD18 without blocking its function, f...

  9. Experimental studies

    Spectral fluence measurements in an adult phantom are reported. A NaI(Tl) probe was used in various locations within the phantom and pulse-height spectra were obtained for seven beam configurations and three generating potentials. Some typical spectra results are presented. A comparison of calculated dose to experimental measurements is presented

  10. Experimental guidelines

    The paper proposes a model experimental design to study the effects of pesticides on particular ecosystem. It takes maize as a model crop and an alternative crop while studying the adverse effects on untargeted arthropods, residues in the soil and other plants. 5 refs, 7 figs

  11. The Relationship between the Antitumor Effect of the IL-12 Gene Therapy and the Expression of Th1 Cytokines in an HPV16-Positive Murine Tumor Model

    Flor García Paz; Vicente Madrid Marina; Ausencio Morales Ortega; Abimelec Santander González; Oscar Peralta Zaragoza; Ana Burguete García; Kirvis Torres Poveda; José Moreno; Juan Alcocer González; Eva Hernandez Marquez; Victor Bermúdez Morales

    2014-01-01

    Objective. The goal of the present study was to investigate the effect of IL-12 expressed in plasmid on the Th1 cytokine profile in an experimental HPV16-positive murine tumor model and the association with the IL-12's antitumor effect. Methods. Mice were injected with BMK-16/myc cells to establish HPV16-positive tumor and then pNGVL3-mIL-12 plasmid; pcDNA3 plasmid or PBS was injected directly into tumor site. The antitumor effect of the treatment was evaluated and the cytokines expression pr...

  12. Gene expression profile of androgen modulated genes in the murine fetal developing lung

    Côté Mélissa

    2010-01-01

    Full Text Available Abstract Background Accumulating evidences suggest that sex affects lung development. Indeed, a higher incidence of respiratory distress syndrome is observed in male compared to female preterm neonates at comparable developmental stage and experimental studies demonstrated an androgen-related delay in male lung maturation. However, the precise mechanisms underlying these deleterious effects of androgens in lung maturation are only partially understood. Methods To build up a better understanding of the effect of androgens on lung development, we analyzed by microarrays the expression of genes showing a sexual difference and those modulated by androgens. Lungs of murine fetuses resulting from a timely mating window of 1 hour were studied at gestational day 17 (GD17 and GD18, corresponding to the period of surge of surfactant production. Using injections of the antiandrogen flutamide to pregnant mice, we hunted for genes in fetal lungs which are transcriptionally modulated by androgens. Results Results revealed that 1844 genes were expressed with a sexual difference at GD17 and 833 at GD18. Many genes were significantly modulated by flutamide: 1597 at GD17 and 1775 at GD18. Datasets were analyzed by using in silico tools for reconstruction of cellular pathways. Between GD17 and GD18, male lungs showed an intensive transcriptional activity of proliferative pathways along with the onset of lung differentiation. Among the genes showing a sex difference or an antiandrogen modulation of their expression, we specifically identified androgen receptor interacting genes, surfactant related genes in particularly those involved in the pathway leading to phospholipid synthesis, and several genes of lung development regulator pathways. Among these latter, some genes related to Shh, FGF, TGF-beta, BMP, and Wnt signaling are modulated by sex and/or antiandrogen treatment. Conclusion Our results show clearly that there is a real delay in lung maturation between

  13. Zika Virus Infection and Development of a Murine Model.

    Shah, Ankit; Kumar, Anil

    2016-08-01

    In view of the recent outbreak of Zika virus (ZIKV), there is an urgent need to investigate the pathogenesis of the symptoms associated with ZIKV infection. Since the first identification of the virus in 1947, the pathologies associated with ZIKV infection were thought to be limited with mild illness that presented fever, rashes, muscle aches, and weakness. However, ZIKV infection has been shown to cause Guillain-Barré Syndrome, and numerous cases of congenital microcephaly in children have been reported when pregnant females were exposed to the virus. The severity and the rate of spread of ZIKV in the last year has drawn alarming interest among researchers to investigate murine models to study viral pathogenesis and develop candidate vaccines. A recent study by Lazear and colleagues, in the May 2016 issue of cell host and microbe, is an effort to study the pathogenesis of contemporary and historical virus strains in various mouse models. PMID:27260223

  14. Isoforms of murine and human serum amyloid P component

    Nybo, Mads; Hackler, R; Kold, B;

    1998-01-01

    affect their number. When the acute-phase response was analysed in three mouse strains, CBA/J and C3H/HeN initially showed seven SAP isoforms in serum and C57BL/6 J three or four. The responses in all three strains peaked at day 2 and were normalized within 14 days. On days 2 and 4, CBA/J and C3H......Isoelectric focusing (IEF) and immunofixation of murine serum amyloid P component (SAP), purified and in serum, showed a distinct and strain-dependent isoform pattern with up to seven bands (pI 5.1-5.7). Neuraminidase treatment caused a shift of the isoforms to more basic pI values, but did not...

  15. Large-scale characterization of the murine cardiac proteome.

    Cosme, Jake; Emili, Andrew; Gramolini, Anthony O

    2013-01-01

    Cardiomyopathies are diseases of the heart that result in impaired cardiac muscle function. This dysfunction can progress to an inability to supply blood to the body. Cardiovascular diseases play a large role in overall global morbidity. Investigating the protein changes in the heart during disease can uncover pathophysiological mechanisms and potential therapeutic targets. Establishing a global protein expression "footprint" can facilitate more targeted studies of diseases of the heart.In the technical review presented here, we present methods to elucidate the heart's proteome through subfractionation of the cellular compartments to reduce sample complexity and improve detection of lower abundant proteins during multidimensional protein identification technology analysis. Analysis of the cytosolic, microsomal, and mitochondrial subproteomes separately in order to characterize the murine cardiac proteome is advantageous by simplifying complex cardiac protein mixtures. In combination with bioinformatic analysis and genome correlation, large-scale protein changes can be identified at the cellular compartment level in this animal model. PMID:23606244

  16. Role of calcium in differentiation of murine erythroleukemia cells

    ZHUDAN; NONGGAOHE; 等

    1993-01-01

    Calcium plays a crucial role in the normal and abnomal cell metabolism.The role of calcium in the differentiation process of murine erythroleukemia cells(MELC)remains controversial.Here,based upon quantitative measurement of fluorescence in single cells,a method was developed to investigate the intracellular free calcium[Ca2+]i concentration and DNA contents simultaneously,by employing the fluorescent probe,fluo-3 acetoxymethyl ester and DNA dye Hoechst 33342.During MELC differentiation.[Ca2+]i concentration incresed.We also demonstrated that calcium ionophore,A23187,enhanced the HMB-induced MELC differentiation,while verapamil,an inhibitor of calcuim uptake,slightly reduced differentiation.These results suggested that an increase in the [Ca2+]i level was an essential step in HMBA-induced MELC differentiation.

  17. Functional expression of murine multidrug resistance in Xenopus laevis oocytes

    Castillo, G.; Vera, J.C.; Rosen, O.M. (Memorial Sloan-Kettering Cancer Research Center, New York, NY (USA)); Yang, Chiaping Huang; Horwitz, S.B. (Albert Einstein College of Medicine, Bronx, NY (USA))

    1990-06-01

    The development of multidrug resistance (MDR) is associated with the overproduction of a plasma membrane glycoprotein, P glycoprotein. Here the authors report the functional expression of a member of the murine MDR family of proteins and show that Xenopus oocytes injected with RNA encoding the mouse mdr1b P glycoprotein develop a MDR-like phenotype. Immunological analysis indicated that oocytes injected with the mdr1b RNA synthesized a protein with the size and immunological characteristics of the mouse mdr1b P glycoprotein. These oocytes exhibited a decreased accumulation of ({sup 3}H)vinblastine and showed an increased capacity to extrude the drug compared to control oocytes not expressing the P glycoprotein. In addition, competition experiments indicated that verapamil, vincristine, daunomycin, and quinidine, but not colchicine, can overcome the rapid drug efflux conferred by the expression of the mouse P glycoprotein.

  18. Statins improve the resolution of established murine venous thrombosis

    Kessinger, Chase W; Kim, Jin Won; Henke, Peter K;

    2015-01-01

    significantly reduced stasis venous thrombus burden by 25% without affecting lipid levels, blood coagulation parameters, or blood cell counts. Statin-driven reductions in VT burden (thrombus mass for stasis thrombi, intravital microscopy thrombus area for non-stasis thrombi) compared similarly to the......-lipid-lowering agents with anti-thrombotic and anti-inflammatory properties-in decreasing thrombus burden and decreasing vein wall injury, mediators of PTS, in established murine stasis and non-stasis chemical-induced venous thrombosis (N = 282 mice). Treatment of mice with daily atorvastatin or rosuvastatin...... therapeutic anticoagulant effects of low molecular weight heparin. Blood from statin-treated mice showed significant reductions in platelet aggregation and clot stability. Statins additionally reduced thrombus plasminogen activator inhibitor-1 (PAI-1), tissue factor, neutrophils, myeloperoxidase, neutrophil...

  19. The immunological relationship between filtrable agent, Salmonella and murine leukosis

    Hamazaki,Yukio

    1977-12-01

    Full Text Available Salmonella typhimurium was invariably isolated from our J strain murine leukosis. Immunization of D103 mice with either inactivated Salmonella typhimurium or the cell-free extract of leukosis inhibited the transplantation of leukosis. The adoptive immunization of D103 mice with spleen cells of Strong A mice immunized with either Salmonella or the cell-free extract of leukosis inhibited the transplantation of leukosis. The addition of either Salmonella or the cell-free extract of leukosis inhibited the migration of macrophages of leukosis spleen in tissue culture. Strong A mice is non-susceptible to J strain leukosis. However, inoculation of neonatal Strong A mice with the cell-free extract of leukosis produced a susceptibility to the transplantation of leukosis. These results suggest that both a filtrable agent and Salmonella typhimurium are present in cells of this leukosis and might be etiologically related to the leukosis.

  20. Macropinocytosis is the Entry Mechanism of Amphotropic Murine Leukemia Virus

    Rasmussen, Izabela; Vilhardt, Frederik

    2015-01-01

    infection. Understanding how pathogens and toxins exploit or divert endocytosis pathways has advanced our understanding of membrane trafficking pathways, which benefits development of new therapeutical schemes and methods of drug delivery. We show here that Murine Leukemia Virus (A-MLV) pseudotyped with the......, or NIH-3T3 cells knocked-down for caveolin expression, was unaffected. Conversely, A-MLV infection of NIH-3T3 and HeLa cells was sensitive to amiloride analogues and actin-depolymerizing drugs that interfere with macropinocytosis. Further manipulation of the actin cytoskeleton through conditional...... amphotropic (expands the host range to many mammalian cells) envelope protein gains entry into host cells by macropinocytosis. Macropinosomes form as large, fluid-filled vacuoles (up to 10 μm) following collapse of cell surface protrusions and membrane scission. We use drugs or introduction of mutant proteins...

  1. The kin17 Protein in Murine Melanoma Cells

    Anelise C. Ramos

    2015-11-01

    Full Text Available kin17 has been described as a protein involved in the processes of DNA replication initiation, DNA recombination, and DNA repair. kin17 has been studied as a potential molecular marker of breast cancer. This work reports the detection and localization of this protein in the murine melanoma cell line B16F10-Nex2 and in two derived subclones with different metastatic potential, B16-8HR and B16-10CR. Nuclear and chromatin-associated protein fractions were analyzed, and kin17 was detected in all fractions, with an elevated concentration observed in the chromatin-associated fraction of the clone with low metastatic potential, suggesting that the kin17 expression level could be a marker of melanoma.

  2. Methylation of inorganic arsenic by murine fetal tissue explants.

    Broka, Derrick; Ditzel, Eric; Quach, Stephanie; Camenisch, Todd D

    2016-07-01

    Although it is generally believed that the developing fetus is principally exposed to inorganic arsenic and the methylated metabolites from the maternal metabolism of arsenic, little is known about whether the developing embryo can autonomously metabolize arsenic. This study investigates inorganic arsenic methylation by murine embryonic organ cultures of the heart, lung, and liver. mRNA for AS3mt, the gene responsible for methylation of arsenic, was detected in all embryonic tissue types studied. In addition, methylated arsenic metabolites were generated by all three tissue types. The fetal liver explants yielded the most methylated arsenic metabolites (∼7% of total arsenic/48 h incubation) while the heart, and lung preparations produced slightly greater than 2% methylated metabolites. With all tissues the methylation proceeded mostly to the dimethylated arsenic species. This has profound implications for understanding arsenic-induced fetal toxicity, particularly if the methylated metabolites are produced autonomously by embryonic tissues. PMID:26446802

  3. Heme Oxygenase-1 Ameliorates Dextran Sulfate Sodium-induced Acute Murine Colitis by Regulating Th17/Treg Cell Balance*

    Zhang, Liya; Zhang, Yanjie; Zhong, Wenwei; Di, Caixia; Lin, Xiaoliang; Xia, Zhenwei

    2014-01-01

    Inflammatory bowel disease (IBD), including ulcerative colitis and Crohn's disease, is a group of autoimmune diseases characterized by nonspecific inflammation in the gastrointestinal tract. Recent investigations suggest that activation of Th17 cells and/or deficiency of regulatory T cells (Treg) is involved in the pathogenesis of IBD. Heme oxygenase (HO)-1 is a protein with a wide range of anti-inflammatory and immune regulatory function, which exerts significantly protective roles in various T cell-mediated diseases. In this study, we aim to explore the immunological regulation of HO-1 in the dextran sulfate sodium-induced model of experimental murine colitis. BALB/c mice were administered 4% dextran sulfate sodium orally; some mice were intraperitoneally pretreated with HO-1 inducer hemin or HO-1 inhibitor stannum protoporphyrin IX. The results show that hemin enhances the colonic expression of HO-1 and significantly ameliorates the symptoms of colitis with improved histological changes, accompanied by a decreased proportion of Th17 cells and increased number of Tregs in mesenteric lymph node and spleen. Moreover, induction of HO-1 down-regulates retinoic acid-related orphan receptor γt expression and IL-17A levels, while promoting Treg-related forkhead box p3 (Foxp3) expression and IL-10 levels in colon. Further study in vitro revealed that up-regulated HO-1 switched the naive T cells to Tregs when cultured under a Th17-inducing environment, which involved in IL-6R blockade. Therefore, HO-1 may exhibit anti-inflammatory activity in the murine model of acute experimental colitis via regulating the balance between Th17 and Treg cells, thus providing a possible novel therapeutic target in IBD. PMID:25112868

  4. Gene expression in IFN-g-activated murine macrophages

    Pereira C.A.

    2004-01-01

    Full Text Available 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.

  5. Deep sequencing of the murine olfactory receptor neuron transcriptome.

    Ninthujah Kanageswaran

    Full Text Available The ability of animals to sense and differentiate among thousands of odorants relies on a large set of olfactory receptors (OR and a multitude of accessory proteins within the olfactory epithelium (OE. ORs and related signaling mechanisms have been the subject of intensive studies over the past years, but our knowledge regarding olfactory processing remains limited. The recent development of next generation sequencing (NGS techniques encouraged us to assess the transcriptome of the murine OE. We analyzed RNA from OEs of female and male adult mice and from fluorescence-activated cell sorting (FACS-sorted olfactory receptor neurons (ORNs obtained from transgenic OMP-GFP mice. The Illumina RNA-Seq protocol was utilized to generate up to 86 million reads per transcriptome. In OE samples, nearly all OR and trace amine-associated receptor (TAAR genes involved in the perception of volatile amines were detectably expressed. Other genes known to participate in olfactory signaling pathways were among the 200 genes with the highest expression levels in the OE. To identify OE-specific genes, we compared olfactory neuron expression profiles with RNA-Seq transcriptome data from different murine tissues. By analyzing different transcript classes, we detected the expression of non-olfactory GPCRs in ORNs and established an expression ranking for GPCRs detected in the OE. We also identified other previously undescribed membrane proteins as potential new players in olfaction. The quantitative and comprehensive transcriptome data provide a virtually complete catalogue of genes expressed in the OE and present a useful tool to uncover candidate genes involved in, for example, olfactory signaling, OR trafficking and recycling, and proliferation.

  6. Co-culture: A quick approach for isolation of street rabies virus in murine neuroblastoma cells

    A. Sasikalaveni

    2015-05-01

    Full Text Available Background: Laboratory detection of rabies in most cases is based on detection of the antigen by fluorescent antibody test, however, in weak positive cases confirmative laboratory diagnosis depends on widely accepted mouse inoculation test. Cell lines like neuroblastoma have been used to isolate the virus with greater success not only to target for diagnosis, but also for molecular studies that determine the epidemiology of the circulating street rabies strains and in studies that look at the efficiency of the developed monoclonal antibodies to neutralize the different rabies strains. Due to the recent issues in obtaining ethical permission for mouse experimentation, and also the passages required in the cell lines to isolate the virus, we report herewith a co-culture protocol using the murine neuroblastoma (MNA cells, which enable quicker isolation of street rabies virus with minimum passages. Objective: This study is not to have an alternative diagnostic assay, but an approach to produce sufficient amount of rabies virus in minimum passages by a co-culture approach in MNA cells. Materials and Methods: The MNA cells are co-cultured by topping the normal cells with infected cells every 48 h and the infectivity was followed up by performing direct fluorescent-antibody test. Results: The co-culture approach results in 100% infectivity and hence the use of live mouse for experimentation could be avoided. Conclusion: Co-culture method provides an alternative for the situations with limited sample volume and for the quicker isolation of virus which warrants the wild type strains without much modification.

  7. Protein-RNA linkage and posttranslational modifications of feline calicivirus and murine norovirus VPg proteins

    Olspert, Allan; Hosmillo, Myra; Chaudhry, Yasmin; Peil, Lauri; Truve, Erkki

    2016-01-01

    Members of the Caliciviridae family of positive sense RNA viruses cause a wide range of diseases in both humans and animals. The detailed characterization of the calicivirus life cycle had been hampered due to the lack of robust cell culture systems and experimental tools for many of the members of the family. However, a number of caliciviruses replicate efficiently in cell culture and have robust reverse genetics systems available, most notably feline calicivirus (FCV) and murine norovirus (MNV). These are therefore widely used as representative members with which to examine the mechanistic details of calicivirus genome translation and replication. The replication of the calicivirus RNA genome occurs via a double-stranded RNA intermediate that is then used as a template for the production of new positive sense viral RNA, which is covalently linked to the virus-encoded protein VPg. The covalent linkage to VPg occurs during genome replication via the nucleotidylylation activity of the viral RNA-dependent RNA polymerase. Using FCV and MNV, we used mass spectrometry-based approach to identify the specific amino acid linked to the 5′ end of the viral nucleic acid. We observed that both VPg proteins are covalently linked to guanosine diphosphate (GDP) moieties via tyrosine positions 24 and 26 for FCV and MNV respectively. These data fit with previous observations indicating that mutations introduced into these specific amino acids are deleterious for viral replication and fail to produce infectious virus. In addition, we also detected serine phosphorylation sites within the FCV VPg protein with positions 80 and 107 found consistently phosphorylated on VPg-linked viral RNA isolated from infected cells. This work provides the first direct experimental characterization of the linkage of infectious calicivirus viral RNA to the VPg protein and highlights that post-translational modifications of VPg may also occur during the viral life cycle. PMID:27375966

  8. Amplification of the murine mdr2 gene and a reconsideration of the structure of the murine mdr gene locus.

    Kirschner, L S

    1995-01-01

    A common feature of cells selected in vitro for the multidrug resistance (MDR) phenotype is the amplification and concomitant overexpression of the mdr genes. In murine macrophage-like J774.2-derived MDR cell lines, there is a good correlation between levels of amplification and expression for the mdr1b gene, but not for the other two gene family members, mdr1a and mdr2. To understand this phenomenon better, a study of the amplification and expression of the mdr2 gene was undertaken. Southern blotting of genomic DNAs from a series of six MDR cell lines revealed that five of these lines had 5'-end amplification of mdr2, whereas only three contained 3'-end amplification. The analysis also suggested the involvement of a recombination hot-spot in this phenomenon. Despite the observation that the ratio between the number of copies of the 5' and 3' ends of the gene differs among cell lines, the ratio of 5' to 3' end transcription of mdr2 was approximately 1 in all cell lines. An analysis of promoter methylation in MDR cell lines demonstrated that this mechanism may play a role in regulating the transcription of mdr2, but not of mdr1b. Long-range mapping of the mdr locus in parental and amplified cell lines suggested that the three mdr genes are oriented in the same direction, and also revealed the presence of a number of rearrangement events. Models for the murine mdr gene locus in wild-type cells and in a cell line containing a rearrangement are presented. PMID:7832992

  9. Infection of Murine Macrophages by Salmonella enterica Serovar Heidelberg Blocks Murine Norovirus Infectivity and Virus-induced Apoptosis.

    Sudhakar S Agnihothram

    Full Text Available Gastroenteritis caused by bacterial and viral pathogens constitutes a major public health threat in the United States accounting for 35% of hospitalizations. In particular, Salmonella enterica and noroviruses cause the majority of gastroenteritis infections, with emergence of sporadic outbreaks and incidence of increased infections. Although mechanisms underlying infections by these pathogens have been individually studied, little is known about the mechanisms regulating co-infection by these pathogens. In this study, we utilized RAW 264.7 murine macrophage cells to investigate the mechanisms governing co-infection with S. enterica serovar Heidelberg and murine norovirus (MNV. We demonstrate that infection of RAW 264.7 cells with S. enterica reduces the replication of MNV, in part by blocking virus entry early in the virus life cycle, and inducing antiviral cytokines later in the infection cycle. In particular, bacterial infection prior to, or during MNV infection affected virus entry, whereas MNV entry remained unaltered when the virus infection preceded bacterial invasion. This block in virus entry resulted in reduced virus replication, with the highest impact on replication observed during conditions of co-infection. In contrast, bacterial replication showed a threefold increase in MNV-infected cells, despite the presence of antibiotic in the medium. Most importantly, we present evidence that the infection of MNV-infected macrophages by S. enterica blocked MNV-induced apoptosis, despite allowing efficient virus replication. This apoptosis blockade was evidenced by reduction in DNA fragmentation and absence of poly-ADP ribose polymerase (PARP, caspase 3 and caspase 9 cleavage events. Our study suggests a novel mechanism of pathogenesis whereby initial co-infection with these pathogens could result in prolonged infection by either of these pathogens or both together.

  10. FLOW CYTOMETRIC COMPARISON OF THE EFFECTS OF TRIALKYTING ON THE MURINE ERYTHROLEUKEMIC CELL

    Cellular effects of exposure to tributyltin (TBT), triethyltin (TET), or trimethyltin (TMT) were investigated by flow cytometry employing the murine erythroleukemic cell (MELC) as a model cellular system. Cell viability was investigated by the carboxyfluorescein diacetate (CFDA) ...

  11. Hematein, a casein kinase II inhibitor, inhibits lung cancer tumor growth in a murine xenograft model

    Hung, Ming-Szu; Xu, Zhidong; Chen, Yu; Smith, Emmanuel; Mao, Jian-Hua; Hsieh, David; Lin, Yu-Ching; Yang, Cheng-Ta; Jablons, David M.; You, Liang

    2013-01-01

    Casein kinase II (CK2) inhibitors suppress cancer cell growth. In this study, we examined the inhibitory effects of a novel CK2 inhibitor, hematein, on tumor growth in a murine xenograft model. We found that in lung cancer cells, hematein inhibited cancer cell growth, Akt/PKB Ser129 phosphorylation, the Wnt/TCF pathway and increased apoptosis. In a murine xenograft model of lung cancer, hematein inhibited tumor growth without significant toxicity to the mice tested. Molecular docking showed t...

  12. Replication of murine cytomegalovirus in lung macrophages: effect of phagocytosis of bacteria.

    Shanley, J D; Pesanti, E L

    1980-01-01

    Murine cytomegalovirus was found to replicate in lung and peritoneal macrophages of both CF-1 and BALB/c mice in vitro. Cytopathic changes typical of cytomegalovirus infection, including intranuclear inclusions, developed within the infected cells and eventually resulted in death of infected macrophages. Viral antigens were demonstrable by indirect immunofluorescence microscopy, and morphologically typical herpesvirus particles were observed in both nuclei and cytoplasm of murine cytomegalovi...

  13. The Immunomodulatory Activity of Jacaric Acid, a Conjugated Linolenic Acid Isomer, on Murine Peritoneal Macrophages

    Liu, Wai Nam; Leung, Kwok Nam

    2015-01-01

    This study aims at demonstrating the immunomodulatory property of jacaric acid, a conjugated linolenic acid (CLNA) isomer that is present in jacaranda seed oil, on murine peritoneal macrophages. Our results showed that jacaric acid exhibited no significant cytotoxicity on the thioglycollate-elicited murine peritoneal macrophages as revealed by the neutral red uptake assay, but markedly increased their cytostatic activity on the T-cell lymphoma MBL-2 cells as measured by the fluorometric CyQua...

  14. ENHANCED ABSORPTION OF MILLIMETER WAVE ENERGY IN MURINE SUBCUTANEOUS BLOOD VESSELS

    Alekseev, Stanislav I.; Ziskin, Marvin C.

    2011-01-01

    The aim of the present study was to determine millimeter wave (MMW) absorption by blood vessels traversing the subcutaneous fat layer of murine skin. Most calculations were performed using the finite-difference time-domain (FDTD) technique. We used two types of models: (1) a rectangular block of multilayer tissue with blood vessels traversing the fat layer and (2) cylindrical models with circular and elliptical cross sections simulating the real geometry of murine limbs. We found that the spe...

  15. α1-giardin based live heterologous vaccine protects against Giardia lamblia infection in a murine model

    Jenikova, Gabriela; Hruz, Petr; Andersson, Karl M.; Tejman-Yarden, Noa; Ferreira, Patricia C. D.; Andersen, Yolanda S.; Davids, Barbara J.; Gillin, Frances D.; Svärd, Staffan G; Curtiss, Roy; Eckmann, Lars

    2011-01-01

    Giardia lamblia is a leading protozoan cause of diarrheal disease worldwide, yet preventive medical strategies are not available. A crude veterinary vaccine has been licensed for cats and dogs, but no defined human vaccine is available. We tested the vaccine potential of three conserved antigens previously identified in human and murine giardiasis, α1-giardin, α-enolase, and ornithine carbamoyl transferase, in a murine model of G. lamblia infection. Live recombinant attenuated Salmonella ente...

  16. 4T1 Murine Mammary Carcinoma Cells Enhance Macrophage-Mediated Innate Inflammatory Responses

    Laurence Madera; Anna Greenshields; Power Coombs, Melanie R.; Hoskin, David W.

    2015-01-01

    Tumor progression and the immune response are intricately linked. While it is known that cancers alter macrophage inflammatory responses to promote tumor progression, little is known regarding how cancers affect macrophage-dependent innate host defense. In this study, murine bone-marrow-derived macrophages (BMDM) were exposed to murine carcinoma-conditioned media prior to assessment of the macrophage inflammatory response. BMDMs exposed to 4T1 mammary carcinoma-conditioned medium demonstrated...

  17. Repeated Bouts of Aerobic Exercise Enhance Regulatory T Cell Responses in a Murine Asthma Model

    Lowder, Thomas; Dugger, Kari; Deshane, Jessy; Estell, Kim; Schwiebert, Lisa M

    2009-01-01

    We have reported previously that moderate intensity aerobic exercise training attenuates airway inflammation in a murine asthma model. Recent studies implicate regulatory T (Treg) cells in decreasing asthma-related airway inflammation; as such, the current study examined the effect of exercise on Treg cell function in a murine asthma model. Mice were sensitized with ovalbumin (OVA) prior to the start of exercise training at a moderate intensity 3× / week for 4 wks; exercise was performed as t...

  18. Assessment of murine lung mechanics outcome measures: alignment with those made in asthmatics

    Walker, Julia K. L.; Kraft, Monica; Fisher, John T

    2013-01-01

    Although asthma is characterized as an inflammatory disease, recent reports highlight the importance of pulmonary physiology outcome measures to the clinical assessment of asthma control and risk of asthma exacerbation. Murine models of allergic inflammatory airway disease have been widely used to gain mechanistic insight into the pathogenesis of asthma; however, several aspects of murine models could benefit from improvement. This review focuses on aligning lung mechanics measures made in mi...

  19. Comparison of histochemical methods for murine eosinophil detection in a RSV vaccine-enhanced inflammation model

    Meyerholz, David K.; Griffin, Michelle A.; Castilow, Elaine M.; Varga, Steven M.

    2009-01-01

    A comparative study of histochemical detection of eosinophils in fixed murine tissue is lacking. Five histochemical methods previously reported for eosinophil detection were quantitatively and qualitatively compared in an established murine RSV vaccine-enhanced inflammation model. Nonspecific neutrophil staining was evaluated in tissue sections of neutrophilic soft tissue lesions and bone marrow from respective animals. Eosinophils had granular red to orange-red cytoplasmic staining, dependin...

  20. Alternate utilization of two regulatory domains within the Moloney murine sarcoma virus long terminal repeat.

    Graves, B J; Eisenberg, S P; Coen, D M; McKnight, S L

    1985-01-01

    The Moloney murine sarcoma virus long terminal repeat (LTR) harbors two distinct positive activators of transcription, namely, a distal signal and an enhancer. In this report we demonstrate that infection by herpes simplex virus (HSV) can markedly affect the utilization of these two Moloney murine sarcoma virus transcription signals. We investigated the HSV-mediated trans-acting effects with two goals in mind: first, to gain insight into LTR function, and second, to probe the mechanisms used ...

  1. Restriction of Human Polyomavirus BK Virus DNA Replication in Murine Cells and Extracts

    Mahon, C.; Liang, B.; Tikhanovich, I.; et al

    2009-01-01

    BK virus (BKV) causes persistent and asymptomatic infections in most humans and is the etiologic agent of polyomavirus-associated nephropathy (PVAN) and other pathologies. Unfortunately, there are no animal models with which to study activation of BKV replication in the human kidney and the accompanying PVAN. Here we report studies of the restriction of BKV replication in murine cells and extracts and the cause(s) of this restriction. Upon infection of murine cells, BKV expressed large T anti...

  2. Fundación del primer bioterio MPF funcional de Colombia Foundation of a functional murine pathogen free animal facility in Colombia

    Ómar Vesga

    2003-02-01

    Full Text Available Cualquier investigación donde se experimente con modelos animales exige que el estado microbiológico y genético de los mismos sea definido desde el principio y verificado periódicamente para garantizar la fiabilidad y certeza de los datos. Buscando producir ratones libres de patógenos murinos en Colombia, se importaron 173 reproductores no emparentados suizos albinos de la cepa ICR que se hospedaron bajo condiciones protocolizadas de aislamiento microbiológico en el Bioterio MPF de la Facultad de Medicina de la Universidad de Antioquia. Tras dos años de funcionamiento ininterrumpido, el control microbiológico demostró que los progenitores mantuvieron inalterada la flora con la cual fueron exportados a Colombia, que sus crías portan flora murina idéntica, y que la misma corresponde a la exigida internacionalmente. Diversas pruebas a la integridad de la barrera microbiológica condujeron a la detección y erradicación de un brote infeccioso por Parvovirus murino. El control reproductivo permitió mantener intacta la heterocigocidad de la cepa, denominada Udea:ICR(CD-1. In order to guarantee the accuracy of the data, experimentation with animal models requires well defined microbiological and genetic statuses with periodical verification. To produce murine pathogen free mice, an outbreeding program was set up by importing 173 non-related Swiss albino ICR breeders that were hosted under strict conditions of microbiological isolation in the animal facilities of the University of Antioquia Medical Faculty. After two years of uninterrupted reproduction, microbiological control demonstrated unaltered murine flora for both parents and offspring, the same with which original breeders were dispatched to Colombia, complying with current international standards. An outbreak of Murine Parvovirus (MPV was timely detected and erradicated thanks to strict testing of microbiological barriers. The breeding program allowed the production of animals free

  3. Experimental Techniques

    Experimental techniques to be used in the new generation of high energy physics are presented. The emphasis is put on the new ATLAS and CMS detectors for the CERN LHC. For the most important elements of these detectors, a description of the underlying physics processes is given, sometimes with reference to comparable detectors used in the past. Some comparative global performances of the two detectors are also given, with reference to benchmark physics processes (detection of the Higgs boson in various mass regions, etc). (author)

  4. Assessment of carbon nanoparticle exposure on murine macrophage function

    Suro-Maldonado, Raquel M.

    There is growing concern about the potential cytotoxicity of nanoparticles. Exposure to respirable ultrafine particles (2.5uM) can adversely affect human health and have been implicated with episodes of increased respiratory diseases such as asthma and allergies. Nanoparticles are of particular interest because of their ability to penetrate into the lung and potentially elicit health effects triggering immune responses. Nanoparticles are structures and devises with length scales in the 1 to 100-nanometer range. Black carbon (BC) nanoparticles have been observed to be products of combustion, especially flame combustion and multi-walled carbon nanotubes (MWCNT) have been shown to be found in both indoor and outdoor air. Furthermore, asbestos, which have been known to cause mesothelioma as well as lung cancer, have been shown to be structurally identical to MWCNTs. The aims of these studies were to examine the effects of carbon nanoparticles on murine macrophage function and clearance mechanisms. Macrophages are immune cells that function as the first line of defense against invading pathogens and are likely to be amongst the first cells affected by nanoparticles. Our research focused on two manufactured nanoparticles, MWCNT and BC. The two were tested against murine-derived macrophages in a chronic contact model. We hypothesized that long-term chronic exposure to carbon nanoparticles would decrease macrophages ability to effectively respond to immunological challenge. Production of nitric oxide (NO), tumor necrosis factor alpha (TNF-alpha), cell surface macrophage; activation markers, reactive oxygen species formation (ROS), and antigen processing and presentation were examined in response to lipopolysaccharide (LPS) following a 144hr exposure to the particulates. Data demonstrated an increase in TNF-alpha, and NO production; a decrease in phagocytosis and antigen processing and presentation; and a decrease in the expression levels of cell surface macrophage

  5. Resistance to cyclopentenylcytosine in murine leukemia L1210 cells.

    Zhang, H; Cooney, D A; Zhang, M H; Ahluwalia, G; Ford, H; Johns, D G

    1993-12-01

    Cyclopentenyl cytosine (CPEC) exhibits oncological activity in murine and human tumor cells and has now entered Phase I clinical trials. Its mode of action as an antitumor agent appears to be inhibition by its triphosphate (CPEC-TP) of CTP synthase, the enzyme which converts UTP to CTP. In an attempt to elucidate the mechanism of resistance to CPEC, a murine leukemia cell line resistant to CPEC (L1210/CPEC) was developed by N-methyl-N-nitro-N-nitrosoguanidine-induced mutagenesis and subsequent selection by cultivation of the L1210 cells in the presence of 2 microM CPEC. Resistant clones were maintained in CPEC-free medium for 6 generations before biochemical studies were performed. The resistant clone selected for further studies was approximately 13,000-fold less sensitive to growth inhibition by CPEC than the parental cells, and the concentration of CPEC required to deplete CTP in the resistant cells was 50-fold higher than in the sensitive cells. A comparison of the kinetic properties of CTP synthase from sensitive and resistant cells indicated alteration in the properties of the enzyme from the latter; the median inhibitory concentration for CPEC-TP increased from 2 to 14 microM, Km for UTP decreased from 126 to 50 microM, and Vmax increased 12-fold from 0.2 to 2.3 nmol/mg/min. Northern blot analyses of polyadenylated RNA from the resistant and sensitive cells indicated a 3-fold increase in transcripts of the CTP synthase gene in the resistant line. Consistent with these alterations in the properties of the enzyme, the resistant cells exhibited significantly expanded CTP and dCTP pools (4- 5-fold) when compared with the sensitive cells. No change was observed, however, in the properties of uridine-cytidine kinase, the enzyme responsible for the initial phosphorylation of CPEC; despite this, however, cellular uptake of CPEC was greatly decreased, and phosphorylation of CPEC and its incorporation into RNA were 10-fold less than in the parental cells. These latter

  6. A role for smoothened during murine lens and cornea development.

    Janet J Y Choi

    Full Text Available Various studies suggest that Hedgehog (Hh signalling plays roles in human and zebrafish ocular development. Recent studies (Kerr et al., Invest Ophthalmol Vis Sci. 2012; 53, 3316-30 showed that conditionally activating Hh signals promotes murine lens epithelial cell proliferation and disrupts fibre differentiation. In this study we examined the expression of the Hh pathway and the requirement for the Smoothened gene in murine lens development. Expression of Hh pathway components in developing lens was examined by RT-PCR, immunofluorescence and in situ hybridisation. The requirement of Smo in lens development was determined by conditional loss-of-function mutations, using LeCre and MLR10 Cre transgenic mice. The phenotype of mutant mice was examined by immunofluorescence for various markers of cell cycle, lens and cornea differentiation. Hh pathway components (Ptch1, Smo, Gli2, Gli3 were detected in lens epithelium from E12.5. Gli2 was particularly localised to mitotic nuclei and, at E13.5, Gli3 exhibited a shift from cytosol to nucleus, suggesting distinct roles for these transcription factors. Conditional deletion of Smo, from ∼E12.5 (MLR10 Cre did not affect ocular development, whereas deletion from ∼E9.5 (LeCre resulted in lens and corneal defects from E14.5. Mutant lenses were smaller and showed normal expression of p57Kip2, c-Maf, E-cadherin and Pax6, reduced expression of FoxE3 and Ptch1 and decreased nuclear Hes1. There was normal G1-S phase but decreased G2-M phase transition at E16.5 and epithelial cell death from E14.5-E16.5. Mutant corneas were thicker due to aberrant migration of Nrp2+ cells from the extraocular mesenchyme, resulting in delayed corneal endothelial but normal epithelial differentiation. These results indicate the Hh pathway is required during a discrete period (E9.5-E12.5 in lens development to regulate lens epithelial cell proliferation, survival and FoxE3 expression. Defective corneal development occurs

  7. Laser-assisted blastocyst dissection and subsequent cultivation of embryonic stem cells in a serum/cell free culture system: applications and preliminary results in a murine model

    Sills Eric

    2006-05-01

    Full Text Available Abstract Background To evaluate embryonic stem cell (ESC harvesting methods with an emphasis on derivation of ESC lines without feeder cells or sera. Using a murine model, laser-assisted blastocyst dissection was performed and compared to conventional immunosurgery to assess a novel laser application for inner cell mass (ICM isolation. Methods Intact blastocysts or isolated ICMs generated in a standard mouse strain were plated in medium with or without serum to compare ESC harvesting efficiency. ESC derivation was also undertaken in a feeder cell-free culture system. Results Although ICM growth and dissociation was comparable irrespective of the media components, an enhanced ESC harvest was observed in our serum-free medium (p Conclusion Achieving successful techniques for human ESC research is fundamentally dependent on preliminary work using experimental animals. In this study, all experimentally developed ESC lines manifested similar features to ESCs obtained from intact blastocysts in standard culture. Cell/sera free murine ESC harvest and propagation are feasible procedures for an embryology laboratory and await refinements for translation to human medical research.

  8. Evidence-Based Translation for the Genomic Responses of Murine Models for the Study of Human Immunity

    Seok, Junhee

    2015-01-01

    Murine models are an essential tool to study human immune responses and related diseases. However, the use of traditional murine models has been challenged by recent systemic surveys that show discordance between human and model immune responses in their gene expression. This is a significant problem in translational biomedical research for human immunity. Here, we describe evidence-based translation (EBT) to improve the analysis of genomic responses of murine models in the translation to hum...

  9. Contribution of Catalase and Superoxide Dismutase to the Intracellular Survival of Clinical Isolates of Staphylococcus aureus in Murine Macrophages

    Das, Debaditya; Bishayi, Biswadev

    2010-01-01

    The present study was performed in order to carefully investigate the interaction of Staphylococcus aureus with murine macrophages and the contribution of catalase and superoxide dismutase in intracellular persistence of Staphylococcus aureus within murine macrophages during in vitro infection. We have reported that Staphylococcus aureus internalized by murine macrophages did not appear to be rapidly killed. Data indicating the contribution of a single catalase and superoxide dismutase in int...

  10. Experimental techniques

    This lecture presents the experimental techniques, developed in the last 10 or 15 years, in order to perform a new class of experiments with exotic nuclei, where the reactions induced by these nuclei allow to get information on their structure. A brief review of the secondary beams production methods will be given, with some examples of facilities in operation or under project. The important developments performed recently on cryogenic targets will be presented. The different detection systems will be reviewed, both the beam detectors before the targets, and the many kind of detectors necessary to detect all outgoing particles after the reaction: magnetic spectrometer for the heavy fragment, detection systems for the target recoil nucleus, γ detectors. Finally, several typical examples of experiments will be detailed, in order to illustrate the use of each detector either alone, or in coincidence with others. (author)

  11. Effects of sodium fluoride on immune response in murine macrophages.

    De la Fuente, Beatriz; Vázquez, Marta; Rocha, René Antonio; Devesa, Vicenta; Vélez, Dinoraz

    2016-08-01

    Excessive fluoride intake may be harmful for health, producing dental and skeletal fluorosis, and effects upon neurobehavioral development. Studies in animals have revealed effects upon the gastrointestinal, renal and reproductive systems. Some of the disorders may be a consequence of immune system alterations. In this study, an in vitro evaluation is made of fluoride immunotoxicity using the RAW 264.7 murine macrophage line over a broad range of concentrations (2.5-75mg/L). The results show that the highest fluoride concentrations used (50-75mg/L) reduce the macrophage population in part as a consequence of the generation of reactive oxygen and/or nitrogen species and consequent redox imbalance, which in turn is accompanied by lipid peroxidation. A decrease in the expression of the antiinflammatory cytokine Il10 is observed from the lowest concentrations (5mg/L). High concentrations (50mg/L) in turn produce a significant increase in the proinflammatory cytokines Il6 and Mip2 from 4h of exposure. In addition, cell phagocytic capacity is seen to decrease at concentrations of ≥20mg/L. These data indicate that fluoride, at high concentrations, may affect macrophages and thus immune system function - particularly with regard to the inflammation autoregulatory processes, in which macrophages play a key role. PMID:26965474

  12. Effects of gold sodium thiomalate on murine spleen cells

    The effects of gold sodium thiomalate (GST) on Balb/c murine spleen cells were investigated using in vitro mitogen blastogenesis techniques. Initial culture of spleen cells in the presence of concanavalin A (Con A) and GST exhibited inhibition of 3H-thymidine uptake due to gold. Spleen cells depleted of monocytes/macrophages by carbonyl iron-ingestion and cultured with Con A and GST demonstrated biphasic effects by gold on thymidine uptake depending upon the Con A and GST concentrations. A final concentration of 2.5 μg/ml con A showed a decreased inhibition of blastogenesis by gold, with macrophage depleted cells as compared to intact spleen cells. While a final concentration of 0.5 μg/ml Con A showed an increased inhibition of blastogenesis by gold with macrophage depleted cells as compared to intact spleen cells. GST was found to be most effective at inhibiting proliferation when present at the initiation of culture. GST was also found to inhibit responsiveness of cells previously incubated with mitogen and its presence in the culture media was required for at least 24-36 hours to produce significant inhibition. These results indicate that the greatest effects of GST on blastogenesis occur during the initial steps of lymphocyte activation and that macrophages may not be the only cell of gold action

  13. Snake venoms components with antitumor activity in murine melanoma cells

    Despite the constant advances in the treatment of cancer, this disease remains one of the main causes of mortality worldwide. So, the development of new treatment modalities is imperative. Snake venom causes a variety of biological effects because they constitute a complex mixture of substances as disintegrins, proteases (serine and metalo), phospholipases A2, L-amino acid oxidases and others. The goal of the present work is to evaluate a anti-tumor activity of some snake venoms fractions. There are several studies of components derived from snake venoms with this kind of activity. After fractionation of snake venoms of the families Viperidae and Elapidae, the fractions were assayed towards murine melanoma cell line B16-F10 and fibroblasts L929. The results showed that the fractions of venom of the snake Notechis ater niger had higher specificity and potential antitumor activity on B16-F10 cell line than the other studied venoms. Since the components of this venom are not explored yet coupled with the potential activity showed in this work, we decided to choose this venom to develop further studies. The cytotoxic fractions were evaluated to identify and characterize the components that showed antitumoral activity. Western blot assays and zymography suggests that these proteins do not belong to the class of metallo and serine proteinases. (author)

  14. Effect of SPG (Sonifilan) immunotherapy and PDT on murine tumor

    PhotoDynamic Therapy of solid tumors is unique in eliciting a strong host immune response unparalleled in other cancer therapies. This immune response is manifested as an acute inflammatory reaction, and can be readily seen as redness and edema around the treated area. Destruction of typical solid tumor cannot be accomplished solely by direct phototoxic action. This was shown to be the case even with drugs more potent in this direct killing effect than Photofrin, the photosensitizer presently used in clinical PDT. Limiting factors seem to be regional insufficiencies in supply of molecular oxygen, needed for generation of phototoxic species. They can be ascribed to the existence of chronically and acute hypoxic tumor regions, oxygen consumption by the photodynamic process, and vascular shutdown induced during PDT. The remaining tumor mass is eradicated by an indirect effect, necrosis induced by destruction of tumor vasculature. Since most events in PDT treated tumor that lead to vascular collapse are, in fact, typical inflammatory manifestations, it was suggested that PDT-induced acute inflammatory reaction actually leads to vascular damage. In a related report characteristics are shown of cellular inflammatory infiltrate in PDT-treated murine tumor. This work examines the effect of combining PDT with immunotherapy, in an attempt to investigate a possibility of amplification of immune reaction to PDT and its direction towards more pervasive destruction of treated tumors. (authors). 6 refs

  15. The murine cardiac 26S proteasome: an organelle awaiting exploration.

    Gomes, Aldrin V; Zong, Chenggong; Edmondson, Ricky D; Berhane, Beniam T; Wang, Guang-Wu; Le, Steven; Young, Glen; Zhang, Jun; Vondriska, Thomas M; Whitelegge, Julian P; Jones, Richard C; Joshua, Irving G; Thyparambil, Sheeno; Pantaleon, Dawn; Qiao, Joe; Loo, Joseph; Ping, Peipei

    2005-06-01

    Multiprotein complexes have been increasingly recognized as essential functional units for a variety of cellular processes, including the protein degradation system. Selective degradation of proteins in eukaryotes is primarily conducted by the ubiquitin proteasome system. The current knowledge base, pertaining to the proteasome complexes in mammalian cells, relies largely upon information gained in the yeast system, where the 26S proteasome is hypothesized to contain a 20S multiprotein core complex and one or two 19S regulatory complexes. To date, the molecular structure of the proteasome system, the proteomic composition of the entire 26S multiprotein complexes, and the specific designated function of individual components within this essential protein degradation system in the heart remain virtually unknown. A functional proteomic approach, employing multidimensional chromatography purification combined with liquid chromatography tandem mass spectrometry and protein chemistry, was utilized to explore the murine cardiac 26S proteasome system. This article presents an overview on the subject of protein degradation in mammalian cells. In addition, this review shares the limited information that has been garnered thus far pertaining to the molecular composition, function, and regulation of this important organelle in the cardiac cells. PMID:16093497

  16. Dystrophic spinal deformities in a neurofibromatosis type 1 murine model.

    Steven D Rhodes

    Full Text Available Despite the high prevalence and significant morbidity of spinal anomalies in neurofibromatosis type 1 (NF1, the pathogenesis of these defects remains largely unknown. Here, we present two murine models: Nf1flox/-;PeriCre and Nf1flox/-;Col.2.3Cre mice, which recapitulate spinal deformities seen in the human disease. Dynamic histomorphometry and microtomographic studies show recalcitrant bone remodeling and distorted bone microarchitecture within the vertebral spine of Nf1flox/-;PeriCre and Nf1flox/-;Col2.3Cre mice, with analogous histological features present in a human patient with dystrophic scoliosis. Intriguingly, 36-60% of Nf1flox/-;PeriCre and Nf1flox/-;Col2.3Cre mice exhibit segmental vertebral fusion anomalies with boney obliteration of the intervertebral disc (IVD. While analogous findings have not yet been reported in the NF1 patient population, we herein present two case reports of IVD defects and interarticular vertebral fusion in patients with NF1. Collectively, these data provide novel insights regarding the pathophysiology of dystrophic spinal anomalies in NF1, and provide impetus for future radiographic analyses of larger patient cohorts to determine whether IVD and vertebral fusion defects may have been previously overlooked or underreported in the NF1 patient population.

  17. Dystrophic spinal deformities in a neurofibromatosis type 1 murine model.

    Rhodes, Steven D; Zhang, Wei; Yang, Dalong; Yang, Hao; Chen, Shi; Wu, Xiaohua; Li, Xiaohong; Yang, Xianlin; Mohammad, Khalid S; Guise, Theresa A; Bergner, Amanda L; Stevenson, David A; Yang, Feng-Chun

    2015-01-01

    Despite the high prevalence and significant morbidity of spinal anomalies in neurofibromatosis type 1 (NF1), the pathogenesis of these defects remains largely unknown. Here, we present two murine models: Nf1flox/-;PeriCre and Nf1flox/-;Col.2.3Cre mice, which recapitulate spinal deformities seen in the human disease. Dynamic histomorphometry and microtomographic studies show recalcitrant bone remodeling and distorted bone microarchitecture within the vertebral spine of Nf1flox/-;PeriCre and Nf1flox/-;Col2.3Cre mice, with analogous histological features present in a human patient with dystrophic scoliosis. Intriguingly, 36-60% of Nf1flox/-;PeriCre and Nf1flox/-;Col2.3Cre mice exhibit segmental vertebral fusion anomalies with boney obliteration of the intervertebral disc (IVD). While analogous findings have not yet been reported in the NF1 patient population, we herein present two case reports of IVD defects and interarticular vertebral fusion in patients with NF1. Collectively, these data provide novel insights regarding the pathophysiology of dystrophic spinal anomalies in NF1, and provide impetus for future radiographic analyses of larger patient cohorts to determine whether IVD and vertebral fusion defects may have been previously overlooked or underreported in the NF1 patient population. PMID:25786243

  18. Human APOBEC3G incorporation into murine leukemia virus particles

    The human APOBEC3G protein exhibits broad antiretroviral activity against a variety of retroviruses. It is packaged into viral particles and executes its antiviral function in the target cell. The packaging of APOBEC3G into different viral particles requires a mechanism that confers this promiscuity. Here, APOBEC3G incorporation into murine leukemia virus (MLV) was studied using retroviral vectors. APOBEC3G uptake did not require either its cytidine deaminase activity or the presence of a retroviral vector genome. Results from immunoprecipitation and co-localization studies of APOBEC3G with a MLV Gag-CFP (cyan fluorescent protein) fusion protein imply an interaction between both proteins. RNase A treatment did not inhibit the co-precipitation of Gag-CFP and APOBEC3G, suggesting that the interaction is RNA independent. Like human immunodeficiency virus (HIV) Gag, the MLV Gag precursor protein appears to interact with APOBEC3G, indicating that Gag contains conserved structures which are used to encapsidate APOBEC3G into different retroviral particles

  19. Suspension culture of Besnoitia caprae by murine macrophage.

    Sadoughifar, R; Namavari, M; Oryan, A

    2015-12-01

    Besnoitia caprae is a tissue cyst-forming protozoan that infects goats and has considerable economic importance in certain regions of Asia and Africa. Murine macrophage J774 cell line was inoculated with tachyzoites of Besnoitia caprae (BC-Pars isolate) collected from mice. A significant growth of tachyzoites was observed in J774. Mice were inoculated with tachyzoites harvested from J774 cell culture. Skin samples from the mice infected with tachyzoites of BC-Pars were PCR positive. One mouse showed alopecia and skin lesions on 45 DPI. Dermal lesions started from around right eye and gradually developed more and more. After euthanasia on 60 DPI, histopathological evaluation of skins around the eye showed necrosis of the epidermis and follicular adnexa with chronic inflammatory cell infiltration. Histopathological sections of their skin showed the presence of necrosis and mononuclear cell infiltration. To the authors' knowledge, this is the first report of successful production of Besnoitia caprae tachyzoites was achieved in vitro by suspension culture technique. Another interesting finding is the report of the alopecia and skin lesions around the eye in mouse that quite similar to lesions of goats due to infection of Besnoitia caprae. PMID:26688623

  20. Cysteine protease activation and apoptosis in Murine norovirus infection

    Ettayebi Khalil

    2009-09-01

    Full Text Available Abstract Background Noroviruses are the leading cause of viral gastroenteritis. Because a suitable in vitro culture system for the human virus has yet to be developed, many basic details of the infection process are unknown. Murine norovirus (MNV serves as a model system for the study of norovirus infection. Recently it was shown that infection of RAW 264.7 cells involved a novel apoptotic pathway involving survivin. Results Using a different set of approaches, the up-regulation of caspases, DNA condensation/fragmentation, and membrane blebbing, all of which are markers of apoptosis, were confirmed. Live cell imaging and activity-based protein profiling showed that activation of caspase-like proteases occurred within two hours of infection, followed by morphological changes to the cells. MNV infection in the presence of caspase inhibitors proceeded via a distinct pathway of rapid cellular necrosis and reduced viral production. Affinity purification of activity-based protein profiling targets and identification by peptide mass fingerprinting showed that the cysteine protease cathepsin B was activated early in infection, establishing this protein as an upstream activator of the intrinsic apoptotic pathway. Conclusion This work adds cathepsin B to the noncanonical programmed cell death induced by MNV, and provides data suggesting that the virus may induce apoptosis to expand the window of time for viral replication. This work also highlights the significant power of activity-based protein profiling in the study of viral pathogenesis.

  1. Murine leukemia virus (MLV replication monitored with fluorescent proteins

    Bittner Alexandra

    2004-12-01

    Full Text Available Abstract Background Cancer gene therapy will benefit from vectors that are able to replicate in tumor tissue and cause a bystander effect. Replication-competent murine leukemia virus (MLV has been described to have potential as cancer therapeutics, however, MLV infection does not cause a cytopathic effect in the infected cell and viral replication can only be studied by immunostaining or measurement of reverse transcriptase activity. Results We inserted the coding sequences for green fluorescent protein (GFP into the proline-rich region (PRR of the ecotropic envelope protein (Env and were able to fluorescently label MLV. This allowed us to directly monitor viral replication and attachment to target cells by flow cytometry. We used this method to study viral replication of recombinant MLVs and split viral genomes, which were generated by replacement of the MLV env gene with the red fluorescent protein (RFP and separately cloning GFP-Env into a retroviral vector. Co-transfection of both plasmids into target cells resulted in the generation of semi-replicative vectors, and the two color labeling allowed to determine the distribution of the individual genomes in the target cells and was indicative for the occurrence of recombination events. Conclusions Fluorescently labeled MLVs are excellent tools for the study of factors that influence viral replication and can be used to optimize MLV-based replication-competent viruses or vectors for gene therapy.

  2. Ectodysplasin A Pathway Contributes to Human and Murine Skin Repair.

    Garcin, Clare L; Huttner, Kenneth M; Kirby, Neil; Schneider, Pascal; Hardman, Matthew J

    2016-05-01

    The highly conserved ectodysplasin A (EDA)/EDA receptor signaling pathway is critical during development for the formation of skin appendages. Mutations in genes encoding components of the EDA pathway disrupt normal appendage development, leading to the human disorder hypohidrotic ectodermal dysplasia. Spontaneous mutations in the murine Eda (Tabby) phenocopy human X-linked hypohidrotic ectodermal dysplasia. Little is known about the role of EDA signaling in adult skin homeostasis or repair. Because wound healing largely mimics the morphogenic events that occur during development, we propose a role for EDA signaling in adult wound repair. Here we report a pronounced delay in healing in Tabby mice, demonstrating a functional role for EDA signaling in adult skin. Moreover, pharmacological activation of the EDA pathway in both Tabby and wild-type mice significantly accelerates healing, influencing multiple processes including re-epithelialization and granulation tissue matrix deposition. Finally, we show that the healing promoting effects of EDA receptor activation are conserved in human skin repair. Thus, targeted manipulation of the EDA/EDA receptor pathway has clear therapeutic potential for the future treatment of human pathological wound healing. PMID:26829034

  3. Immunodetection of osteoadherin in murine tooth extracellular matrices.

    Couble, Marie-Lise; Bleicher, Françoise; Farges, Jean-Christophe; Peyrol, Simone; Lucchini, Marion; Magloire, Henry; Staquet, Marie-Jeanne

    2004-01-01

    An antiserum was generated from synthetic peptides highly conserved between different mammalian species to immunolocalise the small leucine-rich proteoglycan osteoadherin (OSAD) in murine teeth. In 19-day-old embryos of rats and mice, a positive staining was found in incisor predentin and alveolar bone surrounding developing incisors and molars. In newborns, OSAD was detected at the tip of the first molar cusp where it accumulated in predentin concomitantly with odontoblast differentiation. In 2-day-old rats and mice, in the first molar, immunostaining revealed positive predentin, enamel matrix close to the apical pole of ameloblasts and a strong signal in dentin. At this stage, OSAD was detected in predentin in the second molar. Ultrastructural immunocytochemistry showed gold particles associated with collagen fibres in predentin and in foci at the dentin mineralisation front. Gold particles were also detected near the secretory pole of ameloblasts where enamel crystallites elongate. No staining was detected in pulp tissue and dental follicle. Restriction of OSAD expression to the extracellular matrix of bone, dentin and enamel suggests a role of this proteoglycan in the organisation of mineralised tissues. PMID:14673660

  4. Immunotherapy of murine bladder cancer by irradiated tumor vaccine

    Lamm, D.L.; Riggs, D.R.; DeHaven, J.I.; Bryner, R.W. (West Virginia Univ. School of Medicine, Morgantown (USA))

    1991-01-01

    This investigation explored the efficacy of irradiated autologous mouse bladder tumor (Ir-MBT2) as an active specific immunotherapeutic agent and as adjuvant therapy with Bacillus Calmette-Guerin (BCG) against a subcutaneously transplanted murine bladder tumor. Tumor incidence was significantly reduced in groups receiving BCG (27%, p less than 0.005) or Ir-MBT2 with BCG (53%, p less than 0.025), compared to control (93%). Survival was significantly improved in groups treated with BCG (100%, p less than 0.005), 10(5) Ir-MBT2 with BCG (53%, p less than 0.01), or 10(7) Ir-MBT2 with BCG (47%, p less than 0.025) compared with control (13%). Surprisingly, Ir-MBT2 consistently reduced the efficacy of BCG alone. Ir-MBT2 alone (10(7)) appeared to enhance tumor growth. Autologous irradiated bladder tumor vaccine, alone or in combination with BCG, displayed no immunotherapeutic advantage. The use of irradiated tumor cell vaccine for bladder cancer therapy may reduce the results achievable with BCG alone.

  5. Immunotherapy of murine bladder cancer by irradiated tumor vaccine

    This investigation explored the efficacy of irradiated autologous mouse bladder tumor (Ir-MBT2) as an active specific immunotherapeutic agent and as adjuvant therapy with Bacillus Calmette-Guerin (BCG) against a subcutaneously transplanted murine bladder tumor. Tumor incidence was significantly reduced in groups receiving BCG (27%, p less than 0.005) or Ir-MBT2 with BCG (53%, p less than 0.025), compared to control (93%). Survival was significantly improved in groups treated with BCG (100%, p less than 0.005), 10(5) Ir-MBT2 with BCG (53%, p less than 0.01), or 10(7) Ir-MBT2 with BCG (47%, p less than 0.025) compared with control (13%). Surprisingly, Ir-MBT2 consistently reduced the efficacy of BCG alone. Ir-MBT2 alone (10(7)) appeared to enhance tumor growth. Autologous irradiated bladder tumor vaccine, alone or in combination with BCG, displayed no immunotherapeutic advantage. The use of irradiated tumor cell vaccine for bladder cancer therapy may reduce the results achievable with BCG alone

  6. Chromosomal mechanisms in murine radiation acute myeloid leukemogenesis

    Chromosome 2 abnormalities, particularly interstitial deletions, characterize murine radiation-induced acute myeloid leukaemias (AMLs). Here, G-band analyses in CBA/H mice of early (1-6 month) post 3 Gy X-radiation events in bone marrow cells in vivo and karyotype evolution in one unusual AML are presented. The early event analysis showed that all irradiated animals carry chromosome 2 abnormalities, that chromosome 2 abnormalities are more frequent than expected and that interstitial deletions are more common in chromosome 2 than in the remainder of the genome. On presentation AML case N122 carried a t(2; 11) terminal translocation which, with passaging, evolved into a del2(C3F3). Therefore two pathways in leukaemogenesis might exist, one deletion-driven, the other terminal tranlocation-driven involving interstitial genes and terminal genes respectively of chromosome 2. As all irradiated individuals carried chromosome 2 abnormalities, the formation of these aberrations does not determine individual leukaemogenic sensitivity as only 20-25% of animals would be expected to develop AML. Similar lines of argument suggest that chromosome 2 abnormalities are necessary but not sufficient for radiation leukaemogenesis in CBA/H nor are they rate limiting in leukaemogenesis. (Author)

  7. Attenuation of murine graft-versus-host reactivity by azathioprine

    Shand, F.L.

    1980-07-01

    The therapeutic effects of azathioprine were evaluated in a murine graft-versus-host (GVH) model. A single high dose (200 mg/kg) of azathioprine, administered to FI recipients 2 to 3 days after the injection of parental spleen cells, abrogated the ensuing GVH reaction. Lower doses of the drug, even when injected over an extended period of time (14 days), were found to be ineffective. However, high doses of azathioprine failed to protect when FI recipients were sublethally irradiated or injected with cyclophosphamide before GVH induction, and even the transfer of syngeneic FI spleen cells immediately after irradiation failed to alter this outcome. Further analysis of the changes that occurred in sublethally irradiated recipients revealed that the protective effect of azathioprine on CBA leads to (CBA x C57BL)FI GVH reaction was totally abrogated by doses of irradiation as low as 200 rad. Further experiments in which death was used as an indicator of GVH disease showed that lethality could not be reversed in sublethally irradiated FI recipients by the transfer of syngeneic FI spleen cells unless approximately 10 days were allowed to elapse between syngeneic reconstitution and GVH induction. Since syngeneic FI cells from spleen, lymph node, or bone marrow all behaved similarly, it seems likely that the increased severity of GVH reaction induced by prior immunosuppression may not be attributable to a simple cell deletion event.

  8. Attenuation of murine graft-versus-host reactivity by azathioprine

    The therapeutic effects of azathioprine were evaluated in a murine graft-versus-host (GVH) model. A single high dose (200 mg/kg) of azathioprine, administered to FI recipients 2 to 3 days after the injection of parental spleen cells, abrogated the ensuing GVH reaction. Lower doses of the drug, even when injected over an extended period of time (14 days), were found to be ineffective. However, high doses of azathioprine failed to protect when FI recipients were sublethally irradiated or injected with cyclophosphamide before GVH induction, and even the transfer of syngeneic FI spleen cells immediately after irradiation failed to alter this outcome. Further analysis of the changes that occurred in sublethally irradiated recipients revealed that the protective effect of azathioprine on CBA leads to (CBA x C57BL)FI GVH reaction was totally abrogated by doses of irradiation as low as 200 rad. Further experiments in which death was used as an indicator of GVH disease showed that lethality could not be reversed in sublethally irradiated FI recipients by the transfer of syngeneic FI spleen cells unless approximately 10 days were allowed to elapse between syngeneic reconstitution and GVH induction. Since syngeneic FI cells from spleen, lymph node, or bone marrow all behaved similarly, it seems likely that the increased severity of GVH reaction induced by prior immunosuppression may not be attributable to a simple cell deletion event

  9. Chromosomal mechanisms in murine radiation acute myeloid leukemogenesis

    Bouffler, S.D.; Breckon, G.; Cox, R. [National Radiological Protection Board, Chilton (United Kingdom)

    1996-04-01

    Chromosome 2 abnormalities, particularly interstitial deletions, characterize murine radiation-induced acute myeloid leukaemias (AMLs). Here, G-band analyses in CBA/H mice of early (1-6 month) post 3 Gy X-radiation events in bone marrow cells in vivo and karyotype evolution in one unusual AML are presented. The early event analysis showed that all irradiated animals carry chromosome 2 abnormalities, that chromosome 2 abnormalities are more frequent than expected and that interstitial deletions are more common in chromosome 2 than in the remainder of the genome. On presentation AML case N122 carried a t(2; 11) terminal translocation which, with passaging, evolved into a del2(C3F3). Therefore two pathways in leukaemogenesis might exist, one deletion-driven, the other terminal tranlocation-driven involving interstitial genes and terminal genes respectively of chromosome 2. As all irradiated individuals carried chromosome 2 abnormalities, the formation of these aberrations does not determine individual leukaemogenic sensitivity as only 20-25% of animals would be expected to develop AML. Similar lines of argument suggest that chromosome 2 abnormalities are necessary but not sufficient for radiation leukaemogenesis in CBA/H nor are they rate limiting in leukaemogenesis. (Author).

  10. Accumulation of murine amyloid-β mimics early Alzheimer's disease.

    Krohn, Markus; Bracke, Alexander; Avchalumov, Yosef; Schumacher, Toni; Hofrichter, Jacqueline; Paarmann, Kristin; Fröhlich, Christina; Lange, Cathleen; Brüning, Thomas; von Bohlen Und Halbach, Oliver; Pahnke, Jens

    2015-08-01

    Amyloidosis mouse models of Alzheimer's disease are generally established by transgenic approaches leading to an overexpression of mutated human genes that are known to be involved in the generation of amyloid-β in Alzheimer's families. Although these models made substantial contributions to the current knowledge about the 'amyloid hypothesis' of Alzheimer's disease, the overproduction of amyloid-β peptides mimics only inherited (familiar) Alzheimer's disease, which accounts for mild cognitive impairment. Using behavioural tests, electrophysiology and morphological analyses, we compared different ABC transporter-deficient animals and found that alterations are most prominent in neprilysin × ABCC1 double-deficient mice. We show that these mice have a reduced probability to survive, show increased anxiety in new environments, and have a reduced working memory performance. Furthermore, we detected morphological changes in the hippocampus and amygdala, e.g. astrogliosis and reduced numbers of synapses, leading to defective long-term potentiation in functional measurements. Compared to human, murine amyloid-β is poorly aggregating, due to changes in three amino acids at N-terminal positions 5, 10, and 13. Interestingly, our findings account for the action of early occurring amyloid-β species/aggregates, i.e. monomers and small amyloid-β oligomers. Thus, neprilysin × ABCC1 double-deficient mice present a new model for early effects of amyloid-β-related mild cognitive impairment that allows investigations without artificial overexpression of inherited Alzheimer's disease genes. PMID:25991605

  11. The acquisition of cytokine responsiveness by murine B cells

    Poudrier, J; Owens, T

    1994-01-01

    The mechanism whereby small resting (high buoyant density) murine B cells are induced to express interleukin-2 receptors (IL-2R) and to respond to IL-2 was addressed by staining with anti-IL-2R alpha and -IL-2R beta monoclonal antibodies (mAb), and using receptor-specific cDNA probes. Resting B...... staining and mRNA were induced by the combination of LPS plus IL-5. LPS+IL-5-treated B cells responded to IL-2 by Ig secretion. This indicates that B cells regulate their responsiveness to IL-2 similarly to T cells, via the combined level of expression of IL-2R beta and IL-2R alpha. The synergy between IL...... cells expressed undetectable levels of both IL-2R alpha and beta chains on their surface and did not respond to IL-2, even at supra-physiological concentrations. Sepharose-coupled, but not streptavidin-cross-linked, plastic-adsorbed or soluble, anti-mu up-regulated the expression of IL-2R alpha and beta...

  12. Cinnarizine and flunarizine as radiation sensitisers in two murine tumours

    The effect of calcium antagonists, cinnarizine and flunarizine on the radiation sensitivity of two murine tumours, RIF-1 and SCCVII/St was investigated. Initial experiments giving the compounds at 50 mg kg-1 i.p. indicated cinnarizine had no effect on cell survival after 20 Gy of X-rays in the RIF-1 sarcoma and only a small effect in the SCCVII/St carcinoma. Flunarizine produced a small radiosensitisation in the RIF-1 tumour and a substantial sensitisation in the SCCVII/St tumour. Subsequent experiments in the SCCVII/St tumour indicated the optimal radiosensitising dose of flurnarizine was ∼5 mg kg-1, although some sensitisation was apparent throughout the range of 0.05-500 mg kg-1. Flunarizine produced a parallel shift in the X-ray dose response curve, equivalent to a 5-fold reduction in hypoxic fraction. In a normal tissue study, 5 mg kg-1 flunarizine did not enhance reduction in white cell counts produced by X-ray doses of 2-8 Gy. (author)

  13. A comparative parasitologic study on Biomphalaria glabrata snail and C3H/He mice infected with human and murine isolates of Schistosoma mansoni derived from Sumidouro, Rio de Janeiro, Brazil

    Nilcéa Freire

    2003-09-01

    Full Text Available Experiments were carried out to analyze the biological characteristics of two sympatric isolates of Schistosoma mansoni derived from humans and murines in a low endemic transmission area (Sumidouro county, state of Rio de Janeiro, Brazil. Sympatric reared-laboratory Biomphalaria glabrata and C3H/He mice were used as experimental hosts. Parameters assessed comprised: precercarial period, infectivity and mortality (snails, prepatent period, infectivity (percentage of cercariae maturation into adult worm and intestinal egg count (mice. The murine isolate showed a shorter precercarial period and higher infectivity than human isolate (p 0.05. These data suggest that both isolates are local sub-populations, providing support for the hypotheses that in a same biotope mixed populations or sub-populations circulate among their main host (human beings and/or rodent as an anfixenous infection.

  14. Pathogenicity of Trichosporon asahii in a murine model of disseminated trichosporonosis

    2008-01-01

    Background In recent years, superficial and deep mycoses caused by trichosporon were occasionally reported. In 2001, we reported the first case of disseminated trichosporonosis caused by Trichosporon asahii (T. asahii) in China. In this study, the pathogenicity of T. asahii was investigated in a murine model of disseminated trichosporonosis. Methods Seventy-five mice were randomly divided into 7 groups. Each group was inoculated with T. asahii, through intradermal, gastrointestinal tract or intravenous injection. The mice in the experimental groups were given an intraperitoneal injection of cyclophosphamide (CY) to induce granulocytopenia. Mice in the therapeutic group were given both liposomal amphotericin B and fluconazole. The main viscera of the mice were examined by means of tissue culture and pathologic sections. Results In the two intravenous inoculation groups, T. asahii was isolated from at least one organ in 10 of the 12 granulocytopenic mice and 2 of the 14 immunocompetent mice. Two of the 7 mice in the granulocytopenia group presented with lesions in the inoculation position, but none of the 30 mice in the granulocytopenia and the control group which were inoculated intradermally or through the gastrointestinal tract had viscera infection. In the therapeutic group, the ratio of consequently dead mice, the number of involved viscera, and the incidence of systemic infection were significantly less than the untreated group. Acute purulent inflammation and granulomatous inflammation were the main pathological changes in the course of the infection. Arthrospores and filaments were found in the focus. Conclusions T. asahii is an opportunistic pathogen that causes cutaneous and visceral infections in immunologically impaired hosts. An immunocompetent host was to be infected by the invading T. asahii. Several organs, namely the liver, lungs, kidneys, spleen and heart, were predisposed. The therapy of combining liposomal amphotericin B with fluconazole can

  15. Cryoprotectant delivery and removal from murine insulinomas at vitrification-relevant concentrations.

    Mukherjee, Indra Neil; Song, Ying C; Sambanis, Athanassios

    2007-08-01

    Development of optimal cryopreservation protocols requires delivery and removal of cryoprotective agents (CPAs) in such a way that negative osmotic and cytotoxic effects on cells are minimized. This is especially true for vitrification, where high CPA concentrations are employed. In this study, we report on the determination of cell membrane permeability parameters for water (L(p)) and solute (P(s)), and on the design and experimental verification of CPA addition and removal protocols at vitrification-relevant concentrations for a murine insulinoma cell line, betaTC-tet cells. Using membrane permeability values and osmotic tolerance limits, mathematical modeling and computer simulations were used to design CPA addition and removal protocols at high concentrations. The cytotoxic effects of CPAs were also evaluated. Cells were able to tolerate the addition and removal of 2.5M dimethyl sulfoxide (DMSO) and 2.5M 1,2 propanediol (PD) in single steps, but required multi-step addition and removal with 3.0M DMSO, 3.0M PD, and a vitrification-relevant concentration of 3.0M DMSO+3.0M PD. Cytotoxicity studies revealed that betaTC-tet cells were able to tolerate the presence of single component 6.0M DMSO and 6.0M PD and to a lesser extent 3.0M DMSO+3.0M PD. These results determine the time and concentration domain of CPA exposure that cells can tolerate and are essential for designing cryopreservation protocols for free cells as well as cells in engineered tissues. PMID:17533114

  16. Novel Methodology for Characterizing Regional Variations in the Material Properties of Murine Aortas.

    Bersi, Matthew R; Bellini, Chiara; Di Achille, Paolo; Humphrey, Jay D; Genovese, Katia; Avril, Stéphane

    2016-07-01

    Many vascular disorders, including aortic aneurysms and dissections, are characterized by localized changes in wall composition and structure. Notwithstanding the importance of histopathologic changes that occur at the microstructural level, macroscopic manifestations ultimately dictate the mechanical functionality and structural integrity of the aortic wall. Understanding structure-function relationships locally is thus critical for gaining increased insight into conditions that render a vessel susceptible to disease or failure. Given the scarcity of human data, mouse models are increasingly useful in this regard. In this paper, we present a novel inverse characterization of regional, nonlinear, anisotropic properties of the murine aorta. Full-field biaxial data are collected using a panoramic-digital image correlation (p-DIC) system. An inverse method, based on the principle of virtual power (PVP), is used to estimate values of material parameters regionally for a microstructurally motivated constitutive relation. We validate our experimental-computational approach by comparing results to those from standard biaxial testing. The results for the nondiseased suprarenal abdominal aorta from apolipoprotein-E null mice reveal material heterogeneities, with significant differences between dorsal and ventral as well as between proximal and distal locations, which may arise in part due to differential perivascular support and localized branches. Overall results were validated for both a membrane and a thick-wall model that delineated medial and adventitial properties. Whereas full-field characterization can be useful in the study of normal arteries, we submit that it will be particularly useful for studying complex lesions such as aneurysms, which can now be pursued with confidence given the present validation. PMID:27210500

  17. Optimizing the time of doxil injection to increase the drug retention in transplanted murine mammary tumors

    Shaojin You

    2010-03-01

    Full Text Available Shaojin You, Lian Zuo, Wei LiExperimental Cancer Therapeutic Laboratory and Histopathology Core, Atlanta Research and Educational Foundation (151F, Atlanta VA Medical Center, Decatur, GA, USAAbstract: Sex hormonal milieus during the female fertility cycle modulate the tumor vascular permeability of breast cancer. It has been proposed that the liposomal formulated doxorubicin (ie, Doxil, given at the menstrual/estrous stage with the predicted highest tumor vascular permeability, allows significantly increased drug retention in the breast tumor. In the current study, syngeneic murine 4T1 mammary tumors were established on the backs of female BALB/c mice and Doxil was administered at particular mouse estrous cycle stages. The results indicated that Doxil administration during certain times in the mouse estrous cycle was crucial for drug retention in 4T1 tumor tissues. Significantly higher drug concentrations were detected in the tumor tissues when Doxil was administered during the diestrus stage, as compared to when the drug injection was given at all other estrous stages. Our study also showed that the tumor-bearing mice exhibited nearly normal rhythmicity of the estrous cycle post drug injection, indicating the feasibility of continual injection of Doxil at the same estrous cycle stage. By using 4T1 cells cultured in vitro, we showed that progesterone (P4 significantly inhibited cell proliferation and the production of six tumor-derived cytokines, eg, sTNF-RI, CXCL-16, GM-CSF, MIP-1α, MIP-1γ, and Flt3-L. Some of these factors have been shown to be vascular modulators in diverse tissues. In this report, we demonstrated that the concentration of P4 in the plasma and/or estrous cycle stage of 4T1 tumor-bearing mice can be used to select the best time for administrating the liposomal anticancer drugs.Keywords: progesterone, menstrual cycle, mouse mammary tumor, Doxil, breast cancer therapy

  18. Cannabidiol increases survival and promotes rescue of cognitive function in a murine model of cerebral malaria.

    Campos, A C; Brant, F; Miranda, A S; Machado, F S; Teixeira, A L

    2015-03-19

    Cerebral malaria (CM) is a severe complication resulting from Plasmodium falciparum infection that might cause permanent neurological deficits. Cannabidiol (CBD) is a nonpsychotomimetic compound of Cannabis sativa with neuroprotective properties. In the present work, we evaluated the effects of CBD in a murine model of CM. Female mice were infected with Plasmodium berghei ANKA (PbA) and treated with CBD (30mg/kg/day - 3 or 7days i.p.) or vehicle. On 5th day-post-infection (dpi), at the peak of the disease), animals were treated with single or repeated doses of Artesunate, an antimalarial drug. All groups were tested for memory impairment (Novel Object Recognition or Morris Water Maze) and anxiety-like behaviors (Open field or elevated plus maze test) in different stages of the disease (at the peak or after the complete clearance of the disease). Th1/Th2 cytokines and neurotrophins (brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF)) were measured in the prefrontal cortex and hippocampus of experimental groups. PbA-infected mice displayed memory deficits and exhibited increase in anxiety-like behaviors on the 5dpi or after the clearance of the parasitemia, effects prevented by CBD treatment. On 5dpi, TNF-α and IL-6 increased in the hippocampus, while only IL-6 increased in the prefrontal cortex. CBD treatment resulted in an increase in BDNF expression in the hippocampus and decreased levels of proinflammatory cytokines in the hippocampus (TNF-α) and prefrontal cortex (IL-6). Our results indicate that CBD exhibits neuroprotective effects in CM model and might be useful as an adjunctive therapy to prevent neurological symptoms following this disease. PMID:25595981

  19. TREATMENT OF RAT HEPATOMA BY LOCALLY INJECTION OF MURINE IL-12 RETROVIRUS PACKAGING CELL

    2000-01-01

    Objective: To investigate the therapeutic effects of the murine IL-12 (mIL-12) retrovirus packaging cell line on hepatoma injected locally. Methods: The retrovirus vector encoding mIL-12 gene was constructed and transfected into packaging cell line PA317. The cells were then used to treat the rats with experimental orthotopic hepatoma at different time. The therapeutic effects, immune functions of the hosts, pathological and toxicological responses were documented. Results: the results showed that the mIL-12 retrovirus packaging cell line could significantly inhibit the growth of the hepatoma cells injected locally to the hepatoma. The early treatment made the rats survive long, while the medium or late stage treatment could prolong the life time of the rats compared with the bland control group or bland vector control group, though the rats did not survive. The number of NK cells and T cells increased significantly in the treatment group. The effects of the early treatment were superior to those of the medium and late stage treatment. Moreover, the transfection of IL-12 gene locally in the hepatoma tissue could make the hepatoma disappear from other liver lobe. This phenomenon demonstrated that IL-12 could activate the immune cells of the host to kill the untransfected tumor cells. This is very important for IL-12 to be used in gene therapy clinically. Meanwhile, the hepatoma would not recur in the rats that had survived more than 2 months from the early treatment after being re-challenged with tumor cells. Conclusion: the results showed that IL-12 gene injected locally in the hepatoma tissue could enhance the anti-tumor immunity of the host.

  20. Pulsed electric fields for burn wound disinfection in a murine model.

    Golberg, Alexander; Broelsch, G Felix; Vecchio, Daniela; Khan, Saiqa; Hamblin, Michael R; Austen, William G; Sheridan, Robert L; Yarmush, Martin L

    2015-01-01

    Emerging bacterial resistance renders many antibiotics ineffective, making alternative strategies of wound disinfection important. Here the authors report on a new, physical burn wound disinfection method: pulsed electric fields (PEFs). High voltage, short PEFs create nonthermal, permanent damage to cell membranes, possibly by irreversible electroporation. In medicine, PEF technology has recently been used for nonthermal ablation of solid tumors. The authors have expanded the spectrum of PEF applications in medicine to burn wound disinfection. A third-degree burn was induced on the dorsal skin of C57BL/6 mice. Immediately after the injury, the burn wound was infected with Acinetobacter baumannii expressing the luxCDABE operon. Thirty minutes after infection, the infected areas were treated with 80 pulses delivered at 500 V/mm, 70 μs, 1 Hz. The authors used bioluminescence to quantify bacteria on skin. Three animals were used for each experimental condition. PEFs were effective in the disinfection of infected burned murine skin. The bacterial load reduction correlated with the number of delivered pulses. Forty pulses of 500 V/mm led to a 2.04 ± 0.29 Log10 reduction in bacterial load; 80 pulses led to the immediate 5.53 ± 0.30 Log10 reduction. Three hours after PEF, the bacterial reduction of the skin treated with 500 V/mm, 80 pulses was 4.91 ± 0.71 Log10. The authors introduce a new method of wound disinfection using high voltage, short PEFs. They believe that PEF technology may represent an important alternative to antibiotics in addressing bacterial contamination of wounds, particularly those contaminated with multidrug-resistant bacteria. PMID:25167374

  1. Experimental music for experimental physics

    Rosaria Marraffino

    2014-01-01

    Using the sonification technique, physicist and composer Domenico Vicinanza paid homage to CERN at its 60th anniversary ceremony. After months of hard work, he turned the CERN Convention and LHC data into music.   Click here to download the full score of the "LHChamber music". Every birthday deserves gifts and CERN’s 60th anniversary was no exception. Two gifts were very special, thanks to the hard work of Domenico Vicinanza, a physicist and composer. He created two experimental pieces by applying the sonification technique to the CERN Convention and to data recorded by the four LHC detectors during Run 1. “This technique allows us to ‘hear’ data using an algorithm that translates numbers or letters into notes. It keeps the same information enclosed in a graph or a document, but has a more aesthetic exposition,” explains Domenico Vicinanza. “The result is meant to be a metaphor for scientific cooperation, in which d...

  2. Collaborative interactions between type 2 innate lymphoid cells and antigen-specific CD4+ Th2 cells exacerbate murine allergic airway diseases with prominent eosinophilia.

    Liu, Bo; Lee, Jee-Boong; Chen, Chun-Yu; Hershey, Gurjit K Khurana; Wang, Yui-Hsi

    2015-04-15

    Type-2 innate lymphoid cells (ILC2s) and the acquired CD4(+) Th2 and Th17 cells contribute to the pathogenesis of experimental asthma; however, their roles in Ag-driven exacerbation of chronic murine allergic airway diseases remain elusive. In this study, we report that repeated intranasal rechallenges with only OVA Ag were sufficient to trigger airway hyperresponsiveness, prominent eosinophilic inflammation, and significantly increased serum OVA-specific IgG1 and IgE in rested mice that previously developed murine allergic airway diseases. The recall response to repeated OVA inoculation preferentially triggered a further increase of lung OVA-specific CD4(+) Th2 cells, whereas CD4(+) Th17 and ILC2 cell numbers remained constant. Furthermore, the acquired CD4(+) Th17 cells in Stat6(-/-)/IL-17-GFP mice, or innate ILC2s in CD4(+) T cell-ablated mice, failed to mount an allergic recall response to OVA Ag. After repeated OVA rechallenge or CD4(+) T cell ablation, the increase or loss of CD4(+) Th2 cells resulted in an enhanced or reduced IL-13 production by lung ILC2s in response to IL-25 and IL-33 stimulation, respectively. In return, ILC2s enhanced Ag-mediated proliferation of cocultured CD4(+) Th2 cells and their cytokine production, and promoted eosinophilic airway inflammation and goblet cell hyperplasia driven by adoptively transferred Ag-specific CD4(+) Th2 cells. Thus, these results suggest that an allergic recall response to recurring Ag exposures preferentially triggers an increase of Ag-specific CD4(+) Th2 cells, which facilitates the collaborative interactions between acquired CD4(+) Th2 cells and innate ILC2s to drive the exacerbation of a murine allergic airway diseases with an eosinophilic phenotype. PMID:25780046

  3. Enhanced detection and study of murine norovirus-1 using a more efficient microglial cell line

    Lu Yuanan

    2009-11-01

    Full Text Available Abstract Background Human Noroviruses are the predominant cause of non-bacterial gastroenteritis worldwide. To facilitate prevention and control, a norovirus isolated from mice can provide a model to understand human noroviruses. To establish optimal viral infectivity conditions for murine noroviruses, several cell lines of hematopoietic lineage, including murine BV-2, RAW 264.7, and TIB, as well as human CHME-5, were tested comparatively for their sensitivity to murine norovirus-1. Results Except for CHME-5, all three murine-derived cell lines were susceptible to MNV infection. Viral infection of these cells was confirmed by RT-PCR. Using both viral plaque and replication assays, BV-2 and RAW 264.7 cells were determined to have comparable sensitivities to MNV-1 infection. Comparisons of cell growth characteristics, general laboratory handling and potential in-field applications suggest the use of BV-2 to be more advantageous. Conclusion Results obtained from these studies demonstrate that an immortalized microglial cell line can support MNV-1 replication and provides a more efficient method to detect and study murine noroviruses, facilitating future investigations using MNV-1 as a model to study, detect, and control Human Norovirus.

  4. Capto MMC mixed-mode chromatography of murine and rabbit antibodies.

    Arakawa, Tsutomu; Kurosawa, Yasunori; Storms, Michael; Maruyama, Toshiaki; Okumura, C J; Kita, Yoshiko

    2016-11-01

    Murine antibodies have weak affinity for Protein-A. Here, we have tested binding of murine monoclonal antibody (mAb) to Protein-A or Protein-A/Protein-G mixture under salting-out conditions. The addition of ammonium sulfate to HEK conditioned medium (CM) expressing murine mAb resulted in complete binding, leading to its elution by low pH or neutral arginine solution. Alternatively, a mixed-mode chromatography using Capto MMC resin was developed as a capture step. Binding of murine mAb occurred at neutral pH. The bound mAb was eluted with a gradient from 0.3 M NaCl to 0.3 M arginine/0.3 M NaCl at pH 7.0. The Capto MMC-purified murine mAb was further purified by hydroxyl apatite chromatography. Similarly, rabbit mAb was processed with some modifications. Binding of rabbit mAb to Capto MMC required a lower pH. Elution of the bound rabbit mAb was achieved by a gradient to 0.3 M NaCl, pH 7.0. PMID:27444249

  5. PCR master mixes harbour murine DNA sequences. Caveat emptor!

    Philip W Tuke

    Full Text Available BACKGROUND: XMRV is the most recently described retrovirus to be found in Man, firstly in patients with prostate cancer (PC and secondly in 67% of patients with chronic fatigue syndrome (CFS and 3.7% of controls. Both disease associations remain contentious. Indeed, a recent publication has concluded that "XMRV is unlikely to be a human pathogen". Subsequently related but different polytropic MLV (pMLV sequences were also reported from the blood of 86.5% of patients with CFS. and 6.8% of controls. Consequently we decided to investigate blood donors for evidence of XMRV/pMLV. METHODOLOGY/PRINCIPAL FINDINGS: Testing of cDNA prepared from the whole blood of 80 random blood donors, generated gag PCR signals from two samples (7C and 9C. These had previously tested negative for XMRV by two other PCR based techniques. To test whether the PCR mix was the source of these sequences 88 replicates of water were amplified using Invitrogen Platinum Taq (IPT and Applied Biosystems Taq Gold LD (ABTG. Four gag sequences (2D, 3F, 7H, 12C were generated with the IPT, a further sequence (12D by ABTG re-amplification of an IPT first round product. Sequence comparisons revealed remarkable similarities between these sequences, endogeous MLVs and the pMLV sequences reported in patients with CFS. CONCLUSIONS/SIGNIFICANCE: Methodologies for the detection of viruses highly homologous to endogenous murine viruses require special caution as the very reagents used in the detection process can be a source of contamination and at a level where it is not immediately apparent. It is suggested that such contamination is likely to explain the apparent presence of pMLV in CFS.

  6. Transcriptomic analysis of murine embryos lacking endogenous retinoic acid signaling.

    Marie Paschaki

    Full Text Available Retinoic acid (RA, an active derivative of the liposoluble vitamin A (retinol, acts as an important signaling molecule during embryonic development, regulating phenomenons as diverse as anterior-posterior axial patterning, forebrain and optic vesicle development, specification of hindbrain rhombomeres, pharyngeal arches and second heart field, somitogenesis, and differentiation of spinal cord neurons. This small molecule directly triggers gene activation by binding to nuclear receptors (RARs, switching them from potential repressors to transcriptional activators. The repertoire of RA-regulated genes in embryonic tissues is poorly characterized. We performed a comparative analysis of the transcriptomes of murine wild-type and Retinaldehyde Dehydrogenase 2 null-mutant (Raldh2 (-/- embryos - unable to synthesize RA from maternally-derived retinol - using Affymetrix DNA microarrays. Transcriptomic changes were analyzed in two embryonic regions: anterior tissues including forebrain and optic vesicle, and posterior (trunk tissues, at early stages preceding the appearance of overt phenotypic abnormalities. Several genes expected to be downregulated under RA deficiency appeared in the transcriptome data (e.g. Emx2, Foxg1 anteriorly, Cdx1, Hoxa1, Rarb posteriorly, whereas reverse-transcriptase-PCR and in situ hybridization performed for additional selected genes validated the changes identified through microarray analysis. Altogether, the affected genes belonged to numerous molecular pathways and cellular/organismal functions, demonstrating the pleiotropic nature of RA-dependent events. In both tissue samples, genes upregulated were more numerous than those downregulated, probably due to feedback regulatory loops. Bioinformatic analyses highlighted groups (clusters of genes displaying similar behaviors in mutant tissues, and biological functions most significantly affected (e.g. mTOR, VEGF, ILK signaling in forebrain tissues; pyrimidine and purine

  7. Nuclear localization of Annexin A7 during murine brain development

    Noegel Angelika A

    2005-04-01

    Full Text Available Abstract Background Annexin A7 is a member of the annexin protein family, which is characterized by its ability to interact with phospholipids in the presence of Ca2+-ions and which is thought to function in Ca2+-homeostasis. Results from mutant mice showed altered Ca2+-wave propagation in astrocytes. As the appearance and distribution of Annexin A7 during brain development has not been investigated so far, we focused on the distribution of Annexin A7 protein during mouse embryogenesis in the developing central nervous system and in the adult mouse brain. Results Annexin A7 is expressed in cells of the developing brain where a change in its subcellular localization from cytoplasm to nucleus was observed. In the adult CNS, the subcellular distribution of Annexin A7 depends on the cell type. By immunohistochemistry analysis Annexin A7 was detected in the cytosol of undifferentiated cells at embryonic days E5–E8. At E11–E15 the protein is still present in the cytosol of cells predominantly located in the ventricular germinative zone surrounding the lateral ventricle. Later on, at embryonic day E16, Annexin A7 in cells of the intermediate and marginal zone of the neopallium translocates to the nucleus. Neuronal cells of all areas in the adult brain present Annexin A7 in the nucleus, whereas glial fibrillary acidic protein (GFAP-positive astrocytes exhibit both, a cytoplasmic and nuclear staining. The presence of nuclear Annexin A7 was confirmed by extraction of the nucleoplasm from isolated nuclei obtained from neuronal and astroglial cell lines. Conclusion We have demonstrated a translocation of Annexin A7 to nuclei of cells in early murine brain development and the presence of Annexin A7 in nuclei of neuronal cells in the adult animal. The role of Annexin A7 in nuclei of differentiating and mature neuronal cells remains elusive.

  8. Expansion of intestinal epithelial stem cells during murine development.

    Jeffrey J Dehmer

    Full Text Available Murine small intestinal crypt development is initiated during the first postnatal week. Soon after formation, overall increases in the number of crypts occurs through a bifurcating process called crypt fission, which is believed to be driven by developmental increases in the number of intestinal stem cells (ISCs. Recent evidence suggests that a heterogeneous population of ISCs exists within the adult intestine. Actively cycling ISCs are labeled by Lgr5, Ascl2 and Olfm4; whereas slowly cycling or quiescent ISC are marked by Bmi1 and mTert. The goal of this study was to correlate the expression of these markers with indirect measures of ISC expansion during development, including quantification of crypt fission and side population (SP sorting. Significant changes were observed in the percent of crypt fission and SP cells consistent with ISC expansion between postnatal day 14 and 21. Quantitative real-time polymerase chain reaction (RT-PCR for the various ISC marker mRNAs demonstrated divergent patterns of expression. mTert surged earliest, during the first week of life as crypts are initially being formed, whereas Lgr5 and Bmi1 peaked on day 14. Olfm4 and Ascl2 had variable expression patterns. To assess the number and location of Lgr5-expressing cells during this period, histologic sections from intestines of Lgr5-EGFP mice were subjected to quantitative analysis. There was attenuated Lgr5-EGFP expression at birth and through the first week of life. Once crypts were formed, the overall number and percent of Lgr5-EGFP positive cells per crypt remain stable throughout development and into adulthood. These data were supported by Lgr5 in situ hybridization in wild-type mice. We conclude that heterogeneous populations of ISCs are expanding as measured by SP sorting and mRNA expression at distinct developmental time points.

  9. Murine fertilized ovum, blastomere and morula cells lacking SP phenotype

    XU; YiXin; HE; ZhiYing; ZHU; HaiYing; CHEN; XueSong; LI; JianXiu; ZHANG; HongXia; PAN; XingHua

    2007-01-01

    In the field of stem cell research, SP (side population) phenotype is used to define the property that cells maintain a high efflux capability for some fluorescent dye, such as Hoechst 33342. Recently, many researches proposed that SP phenotype is a phenotype shared by some stem cells and some progenitor cells, and that SP phenotype is regarded as a candidate purification marker for stem cells. In this research, murine fertilized ova (including conjugate and single nucleus fertilized ova), 2-cell stage and 8-cell stage blastomeres, morulas and blastocysts were isolated and directly stained by Hoechst 33342 dye. The results show that fertilized ovum, blastomere and morula cells do not demonstrate any ability to efflux the dye. However, the inner cell mass (ICM) cells of blastocyst exhibit SP phenotype, which is consistent with the result of embryonic stem cells (ESCs) in vitro. These results indicate that the SP phenotype of ICM-derived ESCs is an intrinsic property and independent of the culture condition in vitro, and that SP phenotype is one of the characteristics of at least some pluripotent stem cells, but is not shared by totipotent stem cells. In addition, the result that the SP phenotype of ICM cells disappeared when the inhibitor verapamil was added into medium implies that the SP phenotype is directly associated with ABCG2. These results suggest that not all the stem cells demonstrate SP phenotype, and that SP phenotype might act as a purification marker for partial stem cells such as some pluripotent embryonic stem cells and multipotent adult stem cells, but not for all stem cells exampled by the totipotent stem cells in the very early stage of mouse embryos.

  10. Cyclophilin D regulates necrosis, but not apoptosis, of murine eosinophils.

    Zhu, Xiang; Hogan, Simon P; Molkentin, Jeffery D; Zimmermann, Nives

    2016-04-15

    Eosinophil degranulation and clusters of free extracellular granules are frequently observed in diverse diseases, including atopic dermatitis, nasal polyposis, and eosinophilic esophagitis. Whether these intact granules are released by necrosis or a biochemically mediated cytolysis remains unknown. Recently, a peptidyl-prolyl isomerase located within the mitochondrial matrix, cyclophilin D (PPIF), was shown to regulate necrotic, but not apoptotic, cell death in vitro in fibroblasts, hepatocytes, and cardiomyocytes. Whether cyclophilin D regulates necrosis in hematopoietic cells such as eosinophils remains unknown. We used PPIF-deficient (Ppif(-/-)) mice to test whether cyclophilin D is required for regulating eosinophil necrosis. PPIF deficiency did not affect eosinophil development or maturation at baseline. After in vitro ionomycin or H2O2 treatment, Ppif(-/-) eosinophils were significantly protected from Ca(2+) overload- or oxidative stress-induced necrosis. Additionally, Ppif(-/-) eosinophils demonstrated significantly decreased necrosis, but not apoptosis, in response to Siglec-F cross-linking, a stimulus associated with eosinophil-mediated processes in vitro and in vivo. When treated with apoptosis inducers, Ppif(+/+) and Ppif(-/-) eosinophils exhibited no significant difference in apoptosis or secondary necrosis. Finally, in a dextran sodium sulfate-induced colitis model, although levels of colitogenic cytokines and eosinophil-selective chemokines were comparable between Ppif(+/+) and Ppif(-/-) mice, the latter exhibited decreased clinical outcomes. This correlated with significantly reduced eosinophil cytolysis in the colon. Collectively, our present studies demonstrate that murine eosinophil necrosis is regulated in vitro and in vivo by cyclophilin D, at least in part, thus providing new insight into the mechanism of eosinophil necrosis and release of free extracellular granules in eosinophil-associated diseases. PMID:26893161

  11. Tim-4 Inhibits NO Generation by Murine Macrophages.

    Li-yun Xu

    Full Text Available T cell immunoglobulin- and mucin-domain-containing molecule-4 (Tim-4 receives much attention as a potentially negative regulator of immune responses. However, its modulation on macrophages has not been fully elucidated so far. This study aimed to identify the role of Tim-4 in nitric oxide (NO modulation.Macrophages were stimulated with 100 ng/ml LPS or 100 U/ml IFN-γ. RT-PCR was performed to detect TIM-4 mRNA expression. Tim-4 blocking antibody and NF-κB inhibitory ligand were involved in the study. NO levels were assayed by Griess reaction. Phosphorylation of NF-κB, Jak2 or Stat1 was verified by western blot.Tim-4 was up-regulated in murine macrophages after interferon-gamma (IFN-γ stimulation. Tim-4 over-expression decreased NO production and inducible nitric oxide synthase (iNOS expression in lipopolysaccharide (LPS or IFN-γ-stimulated macrophages. Consistently, Tim-4 blockade promoted LPS or IFN-γ-induced NO secretion and iNOS expression. Tim-4 over-expression decreased LPS-induced nuclear factor kappa B (NF-κB p65 phosphorylation in macrophages, which was abrogated by NF-κB inhibitory ligand. On the contrary, Tim-4 blocking increased LPS-induced NF-κB signaling, which was also abrogated by NF-κB inhibition. In addition, Tim-4 blockade promoted Jak2 and Stat1 phosphorylation in IFN-γ stimulated macrophages.These results indicate that Tim-4 is involved in negative regulation of NO production in macrophages, suggesting the critical role of Tim-4 in immune related diseases.

  12. Comparing human norovirus surrogates: murine norovirus and Tulane virus.

    Hirneisen, Kirsten A; Kniel, Kalmia E

    2013-01-01

    Viral surrogates are widely used by researchers to predict human norovirus behavior. Murine norovirus (MNV) is currently accepted as the best surrogate and is assumed to mimic the survival and inactivation of human noroviruses. Recently, a new calicivirus, the Tulane virus (TV), was discovered, and its potential as a human norovirus surrogate is being explored. This study aimed to compare the behavior of the two potential surrogates under varying treatments of pH (2.0 to 10.0), chlorine (0.2 to 2,000 ppm), heat (50 to 75°C), and survival in tap water at room (20°C) and refrigeration (4°C) temperatures for up to 30 days. Viral infectivity was determined by the plaque assay for both MNV and TV. There was no significant difference between the inactivation of MNV and TV in all heat treatments, and for both MNV and TV survival in tap water at 20°C over 30 days. At 4°C, MNV remained infectious over 30 days at a titer of approximately 5 log PFU/ml, whereas TV titers decreased significantly by 5 days. MNV was more pH stable, as TV titers were reduced significantly at pH 2.0, 9.0, and 10.0, as compared with pH 7.0, whereas MNV titers were only significantly reduced at pH 10.0. After chlorine treatment, there was no significant difference in virus with the exception of at 2 ppm, where TV decreased significantly compared with MNV. Compared with TV, MNV is likely a better surrogate for human noroviruses, as MNV persisted over a wider range of pH values, at 2 ppm of chlorine, and without a loss of titer at 4°C. PMID:23317870

  13. Induction and treatment of anergy in murine leprosy.

    Juarez-Ortega, Mario; Hernandez, Víctor G; Arce-Paredes, Patricia; Villanueva, Enrique B; Aguilar-Santelises, Miguel; Rojas-Espinosa, Oscar

    2015-02-01

    Leprosy is a disease consisting of a spectrum of clinical, bacteriological, histopathological and immunological manifestations. Tuberculoid leprosy is frequently recognized as the benign polar form of the disease, while lepromatous leprosy is regarded as the malignant form. The different forms of leprosy depend on the genetic and immunological characteristics of the patient and on the characteristics of the leprosy bacillus. The malignant manifestations of lepromatous leprosy result from the mycobacterial-specific anergy that develops in this form of the disease. Using murine leprosy as a model of anergy in this study, we first induced the development of anergy to Mycobacterium lepraemurium (MLM) in mice and then attempted to reverse it by the administration of dialysable leucocyte extracts (DLE) prepared from healthy (HLT), BCG-inoculated and MLM-inoculated mice. Mice inoculated with either MLM or BCG developed a robust cell-mediated immune response (CMI) that was temporary in the MLM-inoculated group and long-lasting in the BCG-inoculated group. DLE were prepared from the spleens of MLM- and BCG-inoculated mice at the peak of CMI. Independent MLM intradermally-inoculated groups were treated every other day with HLT-DLE, BCG-DLE or MLM-DLE, and the effect was documented for 98 days. DLE administered at a dose of 1.0 U (1 × 10(6) splenocytes) did not affect the evolution of leprosy, while DLE given at a dose of 0.1 U showed beneficial effects regardless of the DLE source. The dose but not the specificity of DLE was the determining factor for reversing anergy. PMID:25529580

  14. Murine macrophage heparanase: inhibition and comparison with metastatic tumor cells

    Circulating macrophages and metastatic tumor cells can penetrate the vascular endothelium and migrate from the circulatory system to extravascular compartments. Both activated murine macrophages and different metastatic tumor cells attach, invade, and penetrate confluent vascular endothelial cell monolayer in vitro, by degrading heparan sulfate proteoglycans in the subendothelial extracellular matrix. The sensitivity of the enzymes from the various sources degrading the heparan sulfate proteoglycan was challenged and compared by a series of inhibitors. Activated macrophages demonstrate a heparanase with an endoglycosidase activity that cleaves from the [35S]O4--labeled heparan sulfate proteoglycans of the extracellular matrix 10 kDa glycosaminoglycan fragments. The degradation of [35S]O4--labeled extracellular matrix proteoglycans by the macrophages' heparanase is significantly inhibited in the presence of heparan sulfate (10μg/ml), arteparon (10μg/ml), and heparin at a concentration of 3 μg/ml. Degradation of this heparan sulfate proteoglycan is a two-step sequential process involving protease activity followed by heparanase activity. B16-BL6 metastatic melanoma cell heparanase, which is also a cell-associated enzyme, was inhibited by heparin to the same extent as the macrophage haparanase. On the other hand, heparanase of the highly metastatic variant (ESb) of a methylcholanthrene-induced T lymphoma, which is an extracellular enzyme released by the cells to the incubation medium, was more sensitive to heparin and arteparon than the macrophages' heparanase. These results may indicate the potential use of heparin or other glycosaminoglycans as specific and differential inhibitors for the formation in certain cases of blood-borne tumor metastasis

  15. Sphingosine kinase 1 in murine dorsal root ganglia

    Dimitra Beroukas

    2015-02-01

    Full Text Available The bioactive sphingolipid, sphingosine 1-phosphate (S1P, is a multifunctional mediator that regulates a multitude of processes such as proliferation and differentiation, immune responses, airway constriction and nociception. S1P is synthesized by two sphingosine kinase isoforms, Sphk1 and Sphk2, which are expressed ubiquitously, but exhibit differential tissue expression patterns among organs. S1P has been shown to be involved in sensory neuron nociceptive signalling. However, the presence and regulation of Sphk expression in sensory neurons under conditions of persistent inflammatory pain are currently unknown. We therefore assessed the expression levels of Sphk in murine dorsal root ganglion (DRG neurons, explored the localisation of Sphk mRNA using In-Situ-Hybridization and used mice with a global null mutation for Sphk1 to investigate the response of sensory neurons in a model of persistent inflammation. Here we showed the expression of both Sphk isoforms in mouse primary sensory neurons. The relative mRNA expression levels for markers of inflammation and nociceptive activity, TNFα and NPY, increased whereas mRNA expression levels for Sphk1 but not Sphk2 decreased in ipsilateral DRG in response to peripheral inflammation. Mice with a global deletion of Sphk1 showed a substantial reduction in Sphk1- but not Sphk2-activity in spinal cord but responded to CFA inflammation in a similar way to control mice, with increased sensitivity to mechanical and thermal stimuli, although the degree of inflammation-induced paw swelling was slightly increased in the Sphk1−/− mice. In summary, Sphk1 mRNA was expressed in virtually all sensory DRG neurons and its expression changed in response to peripheral inflammation. However, deficiency of Sphk1did not impact on the inflammation-dependent changes in the expression of pro-inflammatory markers in DRGs, nor did it significantly change nocifensive behaviour.

  16. Human Leucocyte Antigen-G (HLA-G) and Its Murine Functional Homolog Qa2 in the Trypanosoma cruzi Infection

    Dias, Fabrício C.; Mendes-Junior, Celso T.; Silva, Maria C.; Tristão, Fabrine S. M.; Dellalibera-Joviliano, Renata; Soares, Edson G.; Menezes, Jean G.; Schmidt, André; Dantas, Roberto O.; Marin-Neto, José A.; Silva, João S.; Donadi, Eduardo A.

    2015-01-01

    Genetic susceptibility factors, parasite strain, and an adequate modulation of the immune system seem to be crucial for disease progression after Trypanosoma cruzi infection. HLA-G and its murine functional homolog Qa2 have well-recognized immunomodulatory properties. We evaluated the HLA-G 3′ untranslated region (3′UTR) polymorphic sites (associated with mRNA stability and target for microRNA binding) and HLA-G tissue expression (heart, colon, and esophagus) in patients presenting Chagas disease, stratified according to the major clinical variants. Further, we investigated the transcriptional levels of Qa2 and other pro- and anti-inflammatory genes in affected mouse tissues during T. cruzi experimental acute and early chronic infection induced by the CL strain. Chagas disease patients exhibited differential HLA-G 3′UTR susceptibility allele/genotype/haplotype patterns, according to the major clinical variant (digestive/cardiac/mixed/indeterminate). HLA-G constitutive expression on cardiac muscle and colonic cells was decreased in Chagasic tissues; however, no difference was observed for Chagasic and non-Chagasic esophagus tissues. The transcriptional levels of Qa2 and other anti and proinflammatory (CTLA-4, PDCD1, IL-10, INF-γ, and NOS-2) genes were induced only during the acute T. cruzi infection in BALB/c and C57BL/6 mice. We present several lines of evidence indicating the role of immunomodulatory genes and molecules in human and experimental T. cruzi infection. PMID:25688175

  17. Human Leucocyte Antigen-G (HLA-G and Its Murine Functional Homolog Qa2 in the Trypanosoma cruzi Infection

    Fabrício C. Dias

    2015-01-01

    Full Text Available Genetic susceptibility factors, parasite strain, and an adequate modulation of the immune system seem to be crucial for disease progression after Trypanosoma cruzi infection. HLA-G and its murine functional homolog Qa2 have well-recognized immunomodulatory properties. We evaluated the HLA-G 3′ untranslated region (3′UTR polymorphic sites (associated with mRNA stability and target for microRNA binding and HLA-G tissue expression (heart, colon, and esophagus in patients presenting Chagas disease, stratified according to the major clinical variants. Further, we investigated the transcriptional levels of Qa2 and other pro- and anti-inflammatory genes in affected mouse tissues during T. cruzi experimental acute and early chronic infection induced by the CL strain. Chagas disease patients exhibited differential HLA-G 3′UTR susceptibility allele/genotype/haplotype patterns, according to the major clinical variant (digestive/cardiac/mixed/indeterminate. HLA-G constitutive expression on cardiac muscle and colonic cells was decreased in Chagasic tissues; however, no difference was observed for Chagasic and non-Chagasic esophagus tissues. The transcriptional levels of Qa2 and other anti and proinflammatory (CTLA-4, PDCD1, IL-10, INF-γ, and NOS-2 genes were induced only during the acute T. cruzi infection in BALB/c and C57BL/6 mice. We present several lines of evidence indicating the role of immunomodulatory genes and molecules in human and experimental T. cruzi infection.

  18. Does the crystal habit modulate the genotoxic potential of silica particles? A cytogenetic evaluation in human and murine cell lines.

    Guidi, P; Nigro, M; Bernardeschi, M; Lucchesi, P; Scarcelli, V; Frenzilli, G

    2015-10-01

    Crystalline silica inhaled from occupational sources has been classified by IARC as carcinogenic to humans; in contrast, for amorphous silica, epidemiological and experimental evidence remains insufficient. The genotoxicity of crystalline silica is still debated because of the inconsistency of experimental results ("variability of silica hazard"), often related to the features of the particle surfaces. We have assessed the role of crystal habit in the genotoxicity of silica powders. Pure quartz (crystalline) and vitreous silica (amorphous), sharing the same surface features, were used in an in vitro study with human pulmonary epithelial (A549) and murine macrophage (RAW264.7) cell lines, representative of occupational and environmental exposures. Genotoxicity was evaluated by the comet and micronucleus assays, and cytotoxicity by the trypan blue method. Cells were treated with silica powders for 4 and 24h. Quartz but not vitreous silica caused cell death and DNA damage in RAW264.7 cells. A549 cells were relatively resistant to both powders. Our results support the view that crystal habit per se plays a pivotal role in modulating the biological responses to silica particles. PMID:26433261

  19. Oxidative stress and acute changes in murine brain tissues after nasal instillation of copper particles with different sizes.

    Liu, Yang; Gao, Yuxi; Liu, Ying; Li, Bai; Chen, Chunying; Wu, Gang

    2014-06-01

    We aim to investigate the biological effects of copper particles on the murine brain and their underlying mechanism after nasal instillation of copper particles. We choose different sizes and different concentrations of copper nanoparticles for mice intranasal use. Within one week, the mice were sacrificed. Pathological lesions of glial cells were detected by immunohistochemical assay. Immunohistochemical assay reveals that glial fibrillary acidic protein (GFAP) increased significantly in all experimental groups, especially in nanocopper groups. The ultrastructure of nerve cells was observed through TEM, whose results show that there were chromatin congregation and mitochondria shrinkage in the olfactory cells, and that there was increase of endoplasmic reticulum and disassociation of endoplasmic reticulum ribosomes in hippocampus, particularly in the nanocopper-groups. Oxidative stress indexes were determined with colorimetric methods. There was no significant increase in the antioxidative enzymes (GPX, GST, SOD) in brain tissues; however, significant increase of malondiadehyde (MDA) contents was only found in the Cu nanoparticle-exposed mice at the high dose of 40 mg per kg body weight. Based on the investigation into the biological effects of copper nanoparticles (23.5 nm) after intranasal instillation to the mice, we have found that copper particles can indeed enter into the olfactory bulb and then the deeper brain. The inhalation of high dose copper nanoparticles can induce severer lesions of brain in the experimental mice. The underlying mechanism of copper nanoparticles causing severe brain damage bears little connection with oxidative stress. PMID:24738425

  20. Continued Administration of Ciliary Neurotrophic Factor Protects Mice from Inflammatory Pathology in Experimental Autoimmune Encephalomyelitis

    Kuhlmann, Tanja; Remington, Leah; Cognet, Isabelle; Bourbonniere, Lyne; Zehntner, Simone; Guilhot, Florence; Herman, Alexandra; Guay-Giroux, Angélique; Antel, Jack P.; Owens, Trevor; Gauchat, Jean-François

    2006-01-01

    Multiple sclerosis is an inflammatory disease of the central nervous system that leads to loss of myelin and oligodendrocytes and damage to axons. We show that daily administration (days 8 to 24) of murine ciliary neurotrophic factor (CNTF), a neurotrophic factor that has been described as a survival and differentiation factor for neurons and oligodendrocytes, significantly ameliorates the clinical course of a mouse model of multiple sclerosis. In the acute phase of experimental autoimmune en...

  1. Monoclonal antibody against Klebsiella capsular polysaccharide reduces severity and hematogenic spread of experimental Klebsiella pneumoniae pneumonia.

    Held, T K; Trautmann, M.; Mielke, M E; Neudeck, H; Cryz, S J; Cross, A S

    1992-01-01

    Klebsiella pneumoniae is an important nosocomial pathogen causing severe pulmonary infections. The majority of clinical Klebsiella isolates produce a high-molecular-weight capsular polysaccharide (CPS) which is one of the dominant virulence factors. In the present study, we examined the potency of a murine immunoglobulin M monoclonal antibody (MAb) with specificity to Klebsiella type 2 CPS to protect rats against experimental Klebsiella pneumonia. The MAb did not prevent the invasion of virul...

  2. Tryptase - PAR2 axis in Experimental Autoimmune Prostatitis, a model for Chronic Pelvic Pain Syndrome

    Roman, Kenny; Done, Joseph D.; Schaeffer, Anthony J.; Murphy, Stephen F.; Thumbikat, Praveen

    2014-01-01

    Chronic prostatitis/Chronic pelvic pain syndrome (CP/CPPS) affects up to 15% of the male population and is characterized by pelvic pain. Mast cells are implicated in the murine experimental autoimmune prostatitis (EAP) model as key to chronic pelvic pain development. The mast cell mediator tryptase-β and its cognate receptor protease-activated receptor 2 (PAR2) are involved in mediating pain in other visceral disease models. Prostatic secretions and urines from CP/CPPS patients were examined ...

  3. Experimental Cerebral Malaria Develops Independently of Caspase Recruitment Domain-Containing Protein 9 Signaling

    Julius Clemence R Hafalla; Burgold, Jan; Dorhoi, Anca; Gross, Olaf; Ruland, Jürgen; Stefan H. E. Kaufmann; Matuschewski, Kai

    2012-01-01

    The outcome of infection depends on multiple layers of immune regulation, with innate immunity playing a decisive role in shaping protection or pathogenic sequelae of acquired immunity. The contribution of pattern recognition receptors and adaptor molecules in immunity to malaria remains poorly understood. Here, we interrogate the role of the caspase recruitment domain-containing protein 9 (CARD9) signaling pathway in the development of experimental cerebral malaria (ECM) using the murine Pla...

  4. Erythropoietin treatment alleviates ultrastructural myelin changes induced by murine cerebral malaria

    Hempel, Casper; Hyttel, Poul; Staalsø, Trine;

    2012-01-01

    , adjunctive therapy, which is not available at present. Previously, erythropoietin (EPO) was reported to significantly improve the survival and outcome in a murine CM model. The study objectives were to assess myelin thickness and ultrastructural morphology in the corpus callosum in murine CM and to adress...... for electron microscopy. Myelin sheaths in the corpus callosum were analysed with transmission electron microscopy and stereology. RESULTS: The infection caused clinical CM, which was counteracted by EPO. The total number of myelinated axons was identical in the four groups and mice with CM did not......, perivascular oedemas and intracerebral haemorrhages. CONCLUSIONS: EPO treatment reduced clinical signs of CM and reduced cerebral pathology. Murine CM does not reduce the general thickness of myelin sheaths in the corpus callosum....

  5. Murine model of otitis media with effusion: immunohistochemical demonstration of IL-1 alpha antigen expression.

    Johnson, M D; Contrino, A; Contrino, J; Maxwell, K; Leonard, G; Kreutzer, D

    1994-09-01

    Recent studies have suggested that cytokines likely play a central role in the formation and maintenance of otitis media with effusion (OME). Currently, there is no immunologically defined animal model for the study of cytokines as they contribute to the formation of OME. In the present study, a murine model of OME, using eustachian tube blockage via an external surgical approach, was developed. The murine model temporal bone histology appears to mimic the histology found in chronic otitis media with effusion in humans. Additionally, using this murine model, interleukin-1 alpha (IL-1 alpha) expression was detected in the middle ear using standard immunohistochemical techniques. IL-1 alpha seemed localized to the epithelial lining of the middle ear as well as 5% to 10% of inflammatory cells. This model should provide the necessary tool to further study the immunologic aspects of OME. PMID:8072363

  6. Cloning of murine BRI3 gene and study on its function for inducing cell death

    2002-01-01

    To understand the molecular mechanism of TNFα effects, the cDNA of murine BRI3 gene was cloned from the total RNA of murine brain endothelial cells (bEnd.3)treated with hTNFα by using the suppression subtractive hybridization (SSH) and the RT-PCR method. The fusion expression vector harbouring BRI3 gene and enhanced green fluorescence protein (EGFP) thus obtained were designated as pEGFP/I3. Then pEGFP/I3 was transiently transfected into L929 cells and the fusion protein EGFP/I3 was localized in cytoplasm. It is found that the expression of EGFP/I3 could induce cell death in L929 cells detected by TUNEL method and flow cytometry. And the overexpression of Bci-2 in L929 cells can block cell death induced by EGFP/I3, indicating that murine BRI3 gene might related to the TNFα mediated cytotoxicity.

  7. Structure of the gene encoding the murine protein kinase CK2 beta subunit

    Boldyreff, B; Issinger, O G

    1995-01-01

    The mouse protein kinase CK2 beta subunit gene (Csnk2b) is composed of seven exons contained within 7874 bp. The exon and intron lengths extend from 76 to 321 and 111 to 1272 bp, respectively. The lengths of the murine coding exons correspond exactly to the lengths of the exons in the human CK2...... beta gene. Both genes contain a first untranslated exon. Also, the promoter regions from the human and murine CK2 beta gene share some common features, e.g., they contain neither a TATA nor a CAAT box, exon 1 is flanked by a cluster of CpG dinucleotides and recognition sequences for the Hpa...... has no counterpart in the murine gene. Hence, regulation of transcription of the CK2 beta gene by the catalytic CK2 alpha subunit as was described by Robitzki et al. (J. Biol. Chem. 268: 5694-5703, 1993) for the human gene cannot be considered a general regulatory mechanism....

  8. Changes in gene-expression during development of the murine molar tooth germ

    2007-01-01

    In a matter of a few days the murine tooth germ develops into a complex, mineralized, structure. Murine 30K microarrays were used to examine gene expression in the mandibular first molar tooth germs isolated at 15.5dpc and at 2DPN. Microarray results were validated using real-time RT-PCR. The results suggested that only 25 genes (3 without known functions) exhibited significantly higher expression at 15.5dpc compared to 2DPN. In contrast, almost 1400 genes exhibited significantly (P

  9. Size-Dependent Accumulation of PEGylated Silane-Coated Magnetic Iron Oxide Nanoparticles in Murine Tumors

    Larsen, Esben Kjær Unmack; Nielsen, T.; Wittenborn, T.;

    2009-01-01

    two distinct size subpopulations of 20 and 40 nm mean diameters with increased phagocytic uptake observed for the 40 nm size range in vitro. MRI detection revealed greater iron accumulation in murine tumors for 40 nm nanoparticles after intravenous injection. The enhanced MRI contrast of the larger......Magnetic nanoparticles (MNP) can be used as contrast-enhancing agents to visualize tumors by magnetic resonance imaging (MRI). Here we describe an easy synthesis method of magnetic nanoparticles coated with polyethylene glycol (PEG) and demonstrate size-dependent accumulation in murine tumors...

  10. Multispectral Imaging of T and B Cells in Murine Spleen and Tumor

    Feng, Zipei; Jensen, Shawn M.; Messenheimer, David J.; Farhad, Mohammed; Neuberger, Michael; Bifulco, Carlo B.

    2016-01-01

    Recent advances in multiplex immunohistochemistry techniques allow for quantitative, spatial identification of multiple immune parameters for enhanced diagnostic and prognostic insight. However, applying such techniques to murine fixed tissues, particularly sensitive epitopes, such as CD4, CD8α, and CD19, has been difficult. We compared different fixation protocols and Ag-retrieval techniques and validated the use of multiplex immunohistochemistry for detection of CD3+CD4+ and CD3+CD8+ T cell subsets in murine spleen and tumor. This allows for enumeration of these T cell subsets within immune environments, as well as the study of their spatial distribution. PMID:26994219

  11. Multispectral Imaging of T and B Cells in Murine Spleen and Tumor.

    Feng, Zipei; Jensen, Shawn M; Messenheimer, David J; Farhad, Mohammed; Neuberger, Michael; Bifulco, Carlo B; Fox, Bernard A

    2016-05-01

    Recent advances in multiplex immunohistochemistry techniques allow for quantitative, spatial identification of multiple immune parameters for enhanced diagnostic and prognostic insight. However, applying such techniques to murine fixed tissues, particularly sensitive epitopes, such as CD4, CD8α, and CD19, has been difficult. We compared different fixation protocols and Ag-retrieval techniques and validated the use of multiplex immunohistochemistry for detection of CD3(+)CD4(+) and CD3(+)CD8(+) T cell subsets in murine spleen and tumor. This allows for enumeration of these T cell subsets within immune environments, as well as the study of their spatial distribution. PMID:26994219

  12. Characterization of mouse cellular deoxyribonucleic acid homologous to Abelson murine leukemia virus-specific sequences.

    Dale, B.; Ozanne, B.

    1981-01-01

    The genome of Abelson murine leukemia virus (A-MuLV) consists of sequences derived from both BALB/c mouse deoxyribonucleic acid and the genome of Moloney murine leukemia virus. Using deoxyribonucleic acid linear intermediates as a source of retroviral deoxyribonucleic acid, we isolated a recombinant plasmid which contained 1.9 kilobases of the 3.5-kilobase mouse-derived sequences found in A-MuLV (A-MuLV-specific sequences). We used this clone, designated pSA-17, as a probe restriction enzyme ...

  13. Evaluation of Antibody Responses Elicited by Immunization of Mice with a Pneumococcal Antigen Genetically Fused to Murine HSP70 and Murine Interleukin-4

    Dennis O. GOR; Salamatu S. MAMBULA

    2006-01-01

    The heat shock (stress) protein HSP70 has been shown to be a potent stimulator of cellular immune responses. In order to determine whether HSP70 has the ability to stimulate antibody responses, we constructed and expressed fusion proteins consisting of murine HSP70 or murine interleukin (IL)-4 covalently linked to a pneumococcal cell wall-associated protein antigen designated PpmA. Immunization of mice with the PpmA-HSP70 fusion protein (PpmA-70) failed to elicit an increased PpmA-specific serum antibody response. In contrast, mice immunized with PpmA fused to IL-4 (PpmA-IL4), or PpmA fused to both IL-4 and HSP70 (PpmA-IL4-70) fusion proteins elicited high levels of PpmA-specific antibody responses.These data suggest that HSP70 has a limited capacity to stimulate immune responses to heterologous antigens in vivo.

  14. EFFECTS OF INTERLEUKIN-4 ON GRANULOCYTE-MACROPHAGE-COLONY FORMATION FROM MURINE BONE MARROW CELLS AND HEMATOPOIETIC RECONSTITUTION FOLLOWING MURINE ALLOGENEIC BONE MARROW TRANSPLANTATION

    朱康儿; KerryAtkinson

    1994-01-01

    We investigated the effects of mouse recombinant IL-4 on hematopoiesis in vitro and in vivo.IL-4 alone was found to be incapable of stimulating colony formation,but it inhibited both IL-3-and GM-CSF-induced colony for-mation by murine hematopoietic progenitor cells.In contrast,colony formation induced by G-CSF was enhanced in the presence of IL-4.We also studied the influence of IL-4 on hematopoietie reconstiution after allogeneic bone marrow transplantation in a murine model,and found that IL-4 and G-CSF was significantly suppressed by IL-4.The combination of IL-4 and GM-CSF caused a significant decrease in the absolute mumber of meutrophils.

  15. Transcriptome analysis of the ependymal barrier during murine neurocysticercosis

    Mishra Pramod

    2012-06-01

    to be upregulated at the protein level using immunofluorescence microcopy. This is important, because these molecules are members of the most significant pathways by IPA analyses. Conclusion Thus, our study indicates that ependymal cells actively express immune mediators and likely contribute to the observed immunopathogenesis during infection. Of particular interest is the major upregulation of antigen presentation pathway-related genes and chemokines/cytokines. This could explain how the ependyma is a prominent source of leukocyte infiltration into ventricles through the disrupted ependymal lining by way of pial vessels present in the internal leptomeninges in murine NCC.

  16. Sgk1 Sensitive Pendrin Expression in Murine Platelets

    Lisann Pelzl

    2013-12-01

    Full Text Available Background: The anion exchanger pendrin (SLC26A4 is required for proper development of the inner ear, and contributes to iodide organification in thyroid glands as well as anion transport in various epithelia, such as airways and renal tubules. SLC26A4 deficiency leads to Pendred syndrome, which is characterized by hearing loss with enlarged vestibular aqueducts and variable hypothyroidism and goiter. Pendrin expression in kidney, heart, lung and thyroid is up-regulated by the mineralocorticoid deoxycorticosterone (DOCA. Platelets express anion exchangers but virtually nothing is known about the molecular identity and regulation of those carriers. Other carriers such as the Na+/H+ exchanger are regulated by the mineralocorticoid-sensitive serum and glucocorticoid inducible kinase SGK1. Methods: The present study utilized i quantitative reverse transcription polymerase chain reaction (RT-qPCR to quantify the transcript levels of Slc26a4 as compared to Gapdh and ii western blotting to assess Slc26a4 protein abundance in murine platelets from gene-targeted mice lacking Sgk1 (sgk1-/- and respective wild type animals (sgk1+/+ treated without or with a subcutaneous injection of 2.5 mg DOCA for 3 h, or in sgk1+/+ platelets with or without in vitro treatment for 1 h with 10 µg/ml DOCA. Results: Slc26a4 was expressed in platelets, and in vitro DOCA treatment increased Slc26a4 mRNA levels in platelets isolated from sgk1+/+ mice. Moreover, in vivo DOCA treatment significantly up-regulated Slc26a4 mRNA levels in platelets isolated from sgk1+/+ but not sgk1-/- mice. An increase in Sgk1 mRNA levels paralleled that of Slc26a4 mRNA levels in platelets of sgk1+/+ mice. In addition, DOCA treatment further increased Slc26a4 protein abundance in platelets isolated from sgk1+/+ mice. Conclusions: Pendrin is expressed in platelets and is presumably regulated by SGK1 and mineralocorticoids.

  17. Sleep and fatigue in mice infected with murine gammaherpesvirus 68.

    Olivadoti, Melissa D; Weinberg, Jason B; Toth, Linda A; Opp, Mark R

    2011-05-01

    Fatigue, a common symptom of many acute and chronic medical conditions, reduces both quality of life and workplace productivity and can be disabling. However, the pathophysiologic mechanisms that underlie fatigue can be difficult to study in human populations due to the patient heterogeneity, the variety of underlying causes and potential triggering events, and an inability to collect samples that may be essential to elucidation of mechanisms (e.g., brain). Although the etiology of chronic fatigue syndrome (CFS) remains elusive, some studies have implicated viral infections, including Epstein-Barr virus (EBV), a human gammaherpesvirus, as a potential factor in the pathogenesis of CFS. Murine gammaherpesvirus 68 (γHV68) is a mouse pathogen that shares many similarities with human γHVs, including EBV. In this study, we use γHV68-infected C57BL/6J mice as a model system for studying the impact of chronic viral infection on sleep-wake behavior, activity patterns, and body temperature profiles. Our data show that γHV68 alters sleep, activity, and temperature in a manner suggestive of fatigue. In mice infected with the highest dose used in this study (40,000plaque forming units), food intake, body weight, wheel running, body temperature, and sleep were normal until approximately 7days after infection. These parameters were significantly altered during days 7 through 11, returned to baseline levels at day 12 after infection, and remained within the normal range for the remainder of the 30-day period after inoculation. At that time, both infected and uninfected mice were injected with lipopolysaccharide (LPS), and their responses monitored. Uninfected mice given LPS developed a modest and transient febrile response during the initial light phase (hours 12 through 24) after injection. In contrast, infected mice developed changes in core body temperatures that persisted for at least 5days. Infected mice showed an initial hypothermia that lasted for approximately 12h

  18. Loss of tumorigenicity of murine colon carcinoma MC38/0 cell line after transduction with a retroviral vector carrying murine IL-12 genes

    Pajtasz-Piasecka, E.; Szyda, A.; Rossowska, J.; Krawczenko, A.; Indrová, Marie; Grabarczyk, P.; Wysocki, P.; Mackiewicz, A.; DuĽ, D.

    2004-01-01

    Roč. 50, č. 1 (2004), s. 7-14. ISSN 0015-5500 Grant ostatní: State Committee for Scientific Research of the Republic of Poland (KBN)(PL) PBZ-KBN 004/PO4/98/5f Institutional research plan: CEZ:AV0Z5052915 Keywords : MC38 murine colon carcinoma * IL-12 transduction * tumorigenicity Subject RIV: EC - Immunology Impact factor: 0.507, year: 2004

  19. Anti-Pseudomonas aeruginosa IgY antibodies augment bacterial clearance in a murine pneumonia model

    Thomsen, K; Christophersen, L; Bjarnsholt, T;

    2016-01-01

    -P. aeruginosa IgY antibodies on bacterial eradication in a murine pneumonia model. METHODS: P. aeruginosa pneumonia was established in Balb/c mice and the effects of prophylactic IgY administration on lung bacteriology, clinical parameters and subsequent inflammation were compared to controls. RESULTS...

  20. Dual Role of Host Par2 in a Murine Model of Spontaneous Metastatic B16 Melanoma

    Olejár, Tomáš; Větvička, D.; Zadinová, M.; Poučková, P.; Kukal, J.; Ježek, Petr; Matěj, R.

    2014-01-01

    Roč. 34, č. 7 (2014), s. 3511-3515. ISSN 0250-7005 R&D Projects: GA ČR(CZ) GAP302/10/0346 Institutional support: RVO:67985823 Keywords : PAR2 * melanoma * metastasis * murine model Subject RIV: EA - Cell Biology Impact factor: 1.826, year: 2014

  1. Transgene stability for three replication-competent murine leukemia virus vectors

    Duch, Mogens R.; Carrasco, Maria L; Hansen, Bettina Dencker;

    2004-01-01

    cassette consisting of an internal ribosome entry site followed by the enhanced green fluorescent protein coding sequence inserted in different configurations into murine leukemia virus genomes. In two of the constructs, the insert was located in the upstream part of the U3 region while in the third...

  2. TRIMELLITIC ANHYDRIDE-INDUCED EOSINOPHILIA IN A MURINE MODEL OF OCCUPATIONAL ASTHMA

    TRIMELLITIC ANHYDRIDE-INDUCED EOSINOPHILIA IN A MURINE MODEL OF OCCUPATIONAL ASTHMA. J F Regal, ME Mohrman, E Boykin and D Sailstad. Dept. of Pharmacology, University of Minnesota, Duluth, MN, USA and NHEERL, ORD, US EPA, RTP, NC, USA.Trimellitic anhydride (TMA) is a small m...

  3. Full-Thickness Thermal Injury Delays Wound Closure in a Murine Model

    Wu, Jesse C.; Rose, Lloyd F.; Christy, Robert J.; Leung, Kai P.; Chan, Rodney K.

    2015-01-01

    Objective: The contemporary treatment of a full-thickness burn consists of early eschar excision followed by immediate closure of the open wound using autologous skin. However, most animal models study burn wound healing with the persistence of the burn eschar. Our goal is to characterize a murine model of burn eschar excision to study wound closure kinetics.

  4. Measles virus-specific murine T cell clones: characterization of fine specificity function.

    P. de Vries (Petra); J.P.M. Versteeg-van Oosten (José); I.K.G. Visser (Ilona); R.S. van Binnendijk (Rob); S.A. Langeveld (Sacha); A.D.M.E. Osterhaus (Ab); F.G.C.M. Uytdehaag (Fons)

    1989-01-01

    textabstractMeasles virus (MV)-specific murine helper T cell clones (Thy-1.2+, CD4+, CD8-) were generated from mice immunized with MV-infected mouse brain homogenate by limiting dilution and in vitro stimulation of spleen cells with UV-inactivated MV Ag. The protein specificity of 7 out of 37 stable

  5. Eukaryotic expression, purification, crystallization and preliminary X-ray analysis of murine Manic Fringe

    Jinek, Martin; Conti, Elena

    2006-01-01

    The catalytic domain of the murine glycosyltransferase Manic Fringe was expressed in insect cells. Removal by site-directed mutagenesis of two N-glycosylation sites present in the protein was essential to obtain crystals that diffracted to 1.8 Å resolution.

  6. Cryptic plasmids in Lactobacillus strains isolated from the murine gastrointestinal tract.

    Lin, J H; Savage, D C

    1985-01-01

    Ten of twenty Lactobacillus strains isolated from the gastrointestinal tracts of animals of several species contained plasmids of 80 to 90 megadaltons or less than 2.6 megadaltons in size, as analyzed by agarose gel electrophoresis. The large plasmids were found only in strains originally isolated from the keratinized epithelium of the murine stomach.

  7. Viral Engineering of Chimeric Antigen Receptor Expression on Murine and Human T Lymphocytes.

    Hammill, Joanne A; Afsahi, Arya; Bramson, Jonathan L; Helsen, Christopher W

    2016-01-01

    The adoptive transfer of a bolus of tumor-specific T lymphocytes into cancer patients is a promising therapeutic strategy. In one approach, tumor specificity is conferred upon T cells via engineering expression of exogenous receptors, such as chimeric antigen receptors (CARs). Here, we describe the generation and production of both murine and human CAR-engineered T lymphocytes using retroviruses. PMID:27581020

  8. Murine muscular dystrophy caused by a mutation in the laminin alpha 2 (Lama2) gene

    Xu, H; Wu, X R; Wewer, U M;

    1994-01-01

    The classic murine muscular dystrophy strain, dy, was first described almost 40 years ago. We have identified the molecular basis of an allele of dy, called dy2J, by detecting a mutation in the laminin alpha 2 chain gene--the first identified mutation in laminin-2. The G to A mutation in a splice...

  9. Murine Typhus and Leptospirosis as Causes of Acute Undifferentiated Fever, Indonesia

    Gasem, M.H.; Wagenaar, J.F.P.; Goris, M.G.A.; Adi, M.S.; Isbandrio, B.B.; Hartskeerl, R A; Rolain, J. M.; D. Raoult; Gorp, van, D.M.

    2009-01-01

    To investigate rickettsioses and leptospirosis among urban residents of Semarang, Indonesia, we tested the blood of 137 patients with fever. Evidence of Rickettsia typhi, the agent of murine typhus, was found in 9 patients. Another 9 patients showed inconclusive serologic results. Thirteen patients received a diagnosis of leptospirosis. No dual infections were detected.

  10. Perforin and Fas in murine gammaherpesvirus-specific CD8(+) T cell control and morbidity

    Topham, D J; Cardin, R C; Christensen, Jan Pravsgaard;

    2001-01-01

    The immune system uses both virus-specific T cells and B cells to control the acute and latent phases of respiratory infection with the murine gammaherpesvirus 68 (gammaHV-68). We sought to further define the important effector mechanisms for CD8(+) T cells. First, depletion of the CD4(+) T cells...

  11. ROLE OF COPPER,ZINC-SUPEROXIDE DISMUTASE IN CATALYZING NITROTYROSINE FORMATION IN MURINE LIVER

    The solely known function of Cu,Zn-superoxide dismutase (SOD1) is to catalyze the dismutation of superoxide anion into hydrogen peroxide. Our objective was to determine if SOD1 catalyzed murine liver protein nitration induced by acetaminophen (APAP) and lipopolysaccharide (LPS). Liver and plasma ...

  12. Complete Genome Sequence of Murine Pneumotropic Virus (Polyomaviridae) Clone pKV(37-1).

    Libbey, Jane E; Fujinami, Robert S

    2016-01-01

    The murine pneumotropic virus genome encoded by the pKV(37-1) clone was sequenced to completion. The regulatory region harbored a mutation not previously reported. The protein coding regions (large and small T antigens, viral proteins 1 to 3) showed multiple regions of high amino acid identity to the human, simian, and bovine polyomaviruses. PMID:27198030

  13. Suppression of Murine Lymphocyte Mitogen Responses by Exopolysaccharide from Capnocytophaga ochracea

    Bolton, Ronald W.; Dyer, John K.

    1983-01-01

    An extracellular polysaccharide was purified from culture supernatants of Capnocytophaga ochracea 25, a gram-negative bacillus associated with human periodontal disease. The extracellular polysaccharide suppressed in vitro mitogenic responses of murine splenic lymphocytes to concanavalin A and lipopolysaccharide. This suppression wad dose dependent, persisted up to 120 h, and was not caused by direct toxicity of the extracellular polysaccharide.

  14. Murine Typhus and Leptospirosis as Causes of Acute Undifferentiated Fever, Indonesia

    M.H. Gasem; J.F.P. Wagenaar; M.G.A. Goris; M.S. Adi; B.B. Isbandrio; R.A. Hartskeerl; J.M. Rolain; D. Raoult; E.C.M. van Gorp

    2009-01-01

    To investigate rickettsioses and leptospirosis among urban residents of Semarang, Indonesia, we tested the blood of 137 patients with fever. Evidence of Rickettsia typhi, the agent of murine typhus, was found in 9 patients. Another 9 patients showed inconclusive serologic results. Thirteen patients

  15. CHEMOIMMUNOTHERAPY OF MURINE LIVER METASTASES WITH 5-FLUOROURACIL IN COMBINATION WITH LIPOSOME-ENCAPSULATED MURAMYL DIPEPTIDE

    DAEMEN, T; DONTJE, BHJ; REGTS, J; SCHERPHOF, GL

    1993-01-01

    The therapeutic effect of a combination of liposomal muramyl dipeptide (MDP) and 5-fluorouracil (5FU) was studied in a murine tumor model of hepatic metastases of the tumor cell line C26, a colon adenocarcinoma. Liposomal MDP (250 mug/kg body wt) and a low, nontoxic, dose of 5FU (10 mg/kg body wt) w

  16. Heterologous murine and bovine IVF using bottlenose dolphin (Tursiops truncatus) spermatozoa.

    Sánchez-Calabuig, M J; de la Fuente, J; Laguna-Barraza, R; Beltrán-Breña, P; Martínez-Nevado, E; Johnston, S D; Rizos, D; Gutiérrez-Adán, A; Pérez-Gutiérrez, J F

    2015-10-01

    Assisted reproductive technologies are of great importance for increasing the genetic diversity in captive animals. The use of bovine or murine oocytes in heterologous IVF provides advantages compared to homologous IVF in nondomestic animals, such as the accessibility to oocytes and the availability of well-developed in vitro maturation systems. The aim of this study was to determine the heterologous IVF parameters using cryopreserved dolphin spermatozoa and zona-intact bovine or murine oocytes and to examine the nuclear chromatin status of the dolphin spermatozoa. All the processes involved in the fertilization including embryo cleavage were observed by confocal microscopy and hybrid embryo formation was confirmed by polymerase chain reaction. Heterologous bovine IVF showed no polyspermy, lower percentages of pronuclear formation, and a lower cleavage rate compared to homologous IVF group (34.8% vs. 89.3%). Heterologous murine IVF showed a lower cleavage rate than homologous IVF (9.6% vs. 77.1%). With respect to dolphin sperm chromatin, it was more stable, i.e. more resistant to EDTA-SDS decondensation than the bovine sperm chromatin. This study revealed the stability of the dolphin sperm chromatin and the ability of the dolphin spermatozoa to penetrate zona-intact bovine and murine oocytes, leading to hybrid embryo formation. PMID:26149074

  17. Involvement of Nitric Oxide on Bothropoides insularis Venom Biological Effects on Murine Macrophages In Vitro.

    Ramon R P P B de Menezes

    Full Text Available Viperidae venom has several local and systemic effects, such as pain, edema, inflammation, kidney failure and coagulopathy. Additionally, bothropic venom and its isolated components directly interfere on cellular metabolism, causing alterations such as cell death and proliferation. Inflammatory cells are particularly involved in pathological envenomation mechanisms due to their capacity of releasing many mediators, such as nitric oxide (NO. NO has many effects on cell viability and it is associated to the development of inflammation and tissue damage caused by Bothrops and Bothropoides venom. Bothropoides insularis is a snake found only in Queimada Grande Island, which has markedly toxic venom. Thus, the aim of this work was to evaluate the biological effects of Bothropoides insularis venom (BiV on RAW 264.7 cells and assess NO involvement. The venom was submitted to colorimetric assays to identify the presence of some enzymatic components. We observed that BiV induced H2O2 production and showed proteolytic and phospholipasic activities. RAW 264.7 murine macrophages were incubated with different concentrations of BiV and then cell viability was assessed by MTT reduction assay after 2, 6, 12 and 24 hours of incubation. A time- and concentration-dependent effect was observed, with a tendency to cell proliferation at lower BiV concentrations and cell death at higher concentrations. The cytotoxic effect was confirmed after lactate dehydrogenase (LDH measurement in the supernatant from the experimental groups. Flow cytometry analyses revealed that necrosis is the main cell death pathway caused by BiV. Also, BiV induced NO release. The inhibition of both proliferative and cytotoxic effects with L-NAME were demonstrated, indicating that NO is important for these effects. Finally, BiV induced an increase in iNOS expression. Altogether, these results demonstrate that B. insularis venom have proliferative and cytotoxic effects on macrophages, with

  18. The effect of BW12C on radiosensitivity and necrosis of murine tissues and tumours

    BW12C is a drug that has the potential to induce normal tissue and tumour hypoxia by binding to haemoglobin, increasing its affinity for oxygen and thereby reducing oxygen availability to tissues. Initial results suggested that BW12C administration caused significant radioprotection of normal tissues and induced tumour necrosis, but variable results have been reported subsequently. This work was carried to extend the range of observations concerning the ability of BW12C to radioprotect normal tissues and tumours and to induce necrosis of tumours of the mouse. BW12C was administered as 70 mg/kg intravenous 15 min before irradiation of jejunum in CBA mice and of foot skin in WHT mice with single doses of 240 kVp X-rays while mice breathed gases of varying oxygen tensions. The radiosensitivities of these tissues were assessed by the crypt survival assay and the acute skin reaction, respectively. The radiosensitivity of CaNT tumours to single fraction irradiation was assessed by the regrowth delay assay following administration of single or multiple does of BW12C at varying times to air-breathing CBA mice. The radiation response was compared to the radiosensitivity of clamped tumours. The effect of BW12C alone on tumours was assessed by regrowth delay and histological examination for necrosis. Single or multiple doses of BW12C did not influence the radiosensitivity of CaNT tumours, although marked radioprotection could be induced by clamping the tumours during irradiation. Multiple doses of BW12C alone led to a slight increase in necrosis of the CaNT tumour but did not alter its growth rate. BW12C alone did not induce necrosis of the murine JT lymphoma. The results shown that BW12C did not have a significant effect as a radioprotective or necrotizing agent in these experimental systems. The reported differences in the radiomodifying effects of BW12C are probably tissue-specific and relate to complex biochemical and physiological interactions. 18 refs., 4 figs

  19. Curcumin Ingestion Inhibits Mastocytosis and Suppresses Intestinal Anaphylaxis in a Murine Model of Food Allergy.

    Shannon R M Kinney

    Full Text Available IgE antibodies and mast cells play critical roles in the establishment of allergic responses to food antigens. Curcumin, the active ingredient of the curry spice turmeric, has anti-inflammatory properties, and thus may have the capacity to regulate Th2 cells and mucosal mast cell function during allergic responses. We assessed whether curcumin ingestion during oral allergen exposure can modulate the development of food allergy using a murine model of ovalbumin (OVA-induced intestinal anaphylaxis. Herein, we demonstrate that frequent ingestion of curcumin during oral OVA exposure inhibits the development of mastocytosis and intestinal anaphylaxis in OVA-challenged allergic mice. Intragastric (i.g. exposure to OVA in sensitized BALB/c mice induced a robust IgE-mediated response accompanied by enhanced OVA-IgE levels, intestinal mastocytosis, elevated serum mMCP-1, and acute diarrhea. In contrast, mice exposed to oral curcumin throughout the experimental regimen appeared to be normal and did not exhibit intense allergic diarrhea or a significant enhancement of OVA-IgE and intestinal mast cell expansion and activation. Furthermore, allergic diarrhea, mast cell activation and expansion, and Th2 responses were also suppressed in mice exposed to curcumin during the OVA-challenge phase alone, despite the presence of elevated levels of OVA-IgE, suggesting that curcumin may have a direct suppressive effect on intestinal mast cell activation and reverse food allergy symptoms in allergen-sensitized individuals. This was confirmed by observations that curcumin attenuated the expansion of both adoptively transferred bone marrow-derived mast cells (BMMCs, and inhibited their survival and activation during cell culture. Finally, the suppression of intestinal anaphylaxis by curcumin was directly linked with the inhibition of NF-κB activation in curcumin-treated allergic mice, and curcumin inhibited the phosphorylation of the p65 subunit of NF-κB in BMMCs. In

  20. Static and dynamic mechanics of the murine lung after intratracheal bleomycin

    Papiris Spyridon

    2011-05-01

    Full Text Available Abstract Background Despite its widespread use in pulmonary fibrosis research, the bleomycin mouse model has not been thoroughly validated from a pulmonary functional standpoint using new technologies. Purpose of this study was to systematically assess the functional alterations induced in murine lungs by fibrogenic agent bleomycin and to compare the forced oscillation technique with quasi-static pressure-volume curves in mice following bleomycin exposure. Methods Single intratracheal injections of saline (50 μL or bleomycin (2 mg/Kg in 50 μL saline were administered to C57BL/6 (n = 40 and Balb/c (n = 32 mice. Injury/fibrosis score, tissue volume density (TVD, collagen content, airway resistance (RN, tissue damping (G and elastance coefficient (H, hysteresivity (η, and area of pressure-volume curve (PV-A were determined after 7 and 21 days (inflammation and fibrosis stage, respectively. Statistical hypothesis testing was performed using one-way ANOVA with LSD post hoc tests. Results Both C57BL/6 and Balb/c mice developed weight loss and lung inflammation after bleomycin. However, only C57BL/6 mice displayed cachexia and fibrosis, evidenced by increased fibrosis score, TVD, and collagen. At day 7, PV-A increased significantly and G and H non-significantly in bleomycin-exposed C57BL/6 mice compared to saline controls and further increase in all parameters was documented at day 21. G and H, but not PV-A, correlated well with the presence of fibrosis based on histology, TVD and collagen. In Balb/c mice, no change in collagen content, histology score, TVD, H and G was noted following bleomycin exposure, yet PV-A increased significantly compared to saline controls. Conclusions Lung dysfunction in the bleomycin model is more pronounced during the fibrosis stage rather than the inflammation stage. Forced oscillation mechanics are accurate indicators of experimental bleomycin-induced lung fibrosis. Quasi-static PV-curves may be more sensitive than

  1. Gamma scintigraphy imaging of murine invasive pulmonary aspergillosis with a 111In-labeled cyclic peptide

    Introduction: Invasive pulmonary aspergillosis (IPA) is a leading cause of infection-associated death in immunosuppressed patients. Early detection and early administration of antifungal therapy are critical factors in improving outcome for patients with IPA. Here, we evaluated the imaging properties of a 111In-labeled cyclic peptide targeted to Aspergillus fumigatus in an immunosuppressed murine model of IPA. Methods: A cyclic peptide c(CGGRLGPFC)-NH2 was labeled with 111In by means of diethylenetriaminepentaacetic acid (DTPA). Two days after intranasal inoculation of 17.5x106 conidia of A. fumigatus, mice were injected 111In-DTPA-c(CGGRLGPFC)-NH2 intravenously. Biodistribution data were obtained at 2 h, and γ-images were acquired at 10 min and 2 h after radiotracer injection. Healthy mice were used as controls. In addition, a group of infected mice were co-injected with the radiotracer and unlabeled c(CGGRLGPFC)-NH2 to evaluate the inhibition of radiotracer's binding to infected lungs. Autoradiographs of lungs from infected and healthy mice were compared with corresponding photographs of transaxial sections of the lung tissues stained for A. fumigatus hyphae. Results: The labeling efficiency was >98%, with specific radioactivity of up to 74 MBq/nmol peptide. Significantly higher uptake of 111In-DTPA-c(CGGRLGPFC)-NH2 was observed in the lungs of mice infected with A. fumigatus than in those of healthy mice (0.37±0.06 %ID/g vs. 0.14±0.02 %ID/g, P=.00044). Simultaneous injection with unlabeled peptide reduced radioactivity in the infected lungs by 41% (P=.0037). Increased radioactivity in the lungs of infected mice was visible in γ images at both 10 min and 2 h after radiotracer injection. Moreover, autoradiography confirmed radiotracer uptake in infected lungs, but not in the lungs of healthy mice or infected mice co-injected with unlabeled peptide. Conclusions: γ-Imaging with 111In-DTPA-c(CGGRLGPFC)-NH2 clearly delineated experimental IPA in mice. Peptides

  2. Respiratory syncytial virus infection in immunocompetent and immunocompromised murine

    JUAN ZHOU; YU XIA CUI; XI QIANG YANG; ZHOU FU; LI PING JIANG; LI JIA WANG

    2006-01-01

    The purpose of this study is to distinguish respiratory syncytial virus (RSV) infection and immunology between immunocompetent and immunocompromised murine and to explore immune mechanism of RSV infection. At various time points after RSV infection of BALB/c mice and nude mice, pulmonary viral titers were assayed, RSV antigen was tested by direct immunofluorescent assay and immunohistochemistry. Pulmonary mRNA expressions of Toll like receptor (TLR)2 and TLR4 were assayed by RT-PCR. CD4+ cells and CD8+ cells in peripheral blood were examined by flow cytometry and plasma total IgE was assayed by ELISA. Leukocytes in bronchoalveolar lavage fluid (BALF) and pulmonary histology were identified to reflect airway inflammation. It was found that RSV titers of both mice peaked on the 3rd day post infection with a much higher level of viral titer in nude mice than in BALB/c mice and a longer viral duration in nude mice (over 9 days post infection) than in BALB/c mice (6 days post infection). RSV infection induced higher viral antigen expression in nude mice (0.267 ±0.045) than in BALB/c mice (0. 168 ± 0.031). RSV infection enhanced pulmonary TLR4 expression of BALB/c mice (51.96% ± 11.34%) and nude mice (48.96% ± 12.35%) compared with each control (34.04% ± 10.06% and 32.37% ± 9.87% respectively). CD4+ peripheral blood cells increased in RSV infected BALB/c mice (66.51% ± 2.09% ) compared with the control BALB/c mice (51.63% ± 5.90% ), and CD4+ cells and CD8+ cells were deficient in nude mice. RSV infection increased plasma total IgE in both mice, and BALB/c mice had a larger amount of IgE on the 7th day post infection (9.02 ng/ml ± 2.90 ng/ml) and on the 14th day post infection (12.76 ng/ml ± 4.15 ng/ml) than corresponding nude mice (3.72 ng/ml ± 1.06 ng/ml and 7.62 ng/ml ± 3.08 ng/ml respectively on the 7th and 14th day post infection). RSV infected nude mice had more severe airway inflammation than infected BALB/c mice. It is concluded that BALB/c mice and

  3. The AOM/DSS murine model for the study of colon carcinogenesis: From pathways to diagnosis and therapy studies

    Mariangela De Robertis

    2011-01-01

    Full Text Available Colorectal cancer (CRC is a major health problem in industrialized countries. Although inflammation-linked carcinogenesis is a well accepted concept and is often observed within the gastrointestinal tract, the underlying mechanisms remain to be elucidated. Inflammation can indeed provide initiating and promoting stimuli and mediators, generating a tumour-prone microenvironment. Many murine models of sporadic and inflammation-related colon carcinogenesis have been developed in the last decade, including chemically induced CRC models, genetically engineered mouse models, and xenoplants. Among the chemically induced CRC models, the combination of a single hit of azoxymethane (AOM with 1 week exposure to the inflammatory agent dextran sodium sulphate (DSS in rodents has proven to dramatically shorten the latency time for induction of CRC and to rapidly recapitulate the aberrant crypt foci-adenoma-carcinoma sequence that occurs in human CRC. Because of its high reproducibility and potency, as well as the simple and affordable mode of application, the AOM/DSS has become an outstanding model for studying colon carcinogenesis and a powerful platform for chemopreventive intervention studies. In this article we highlight the histopathological and molecular features and describe the principal genetic and epigenetic alterations and inflammatory pathways involved in carcinogenesis in AOM/DSS-treated mice; we also present a general overview of recent experimental applications and preclinical testing of novel therapeutics in the AOM/DSS model.

  4. Effect of pecan phenolics on the release of nitric oxide from murine RAW 264.7 macrophage cells.

    Robbins, Katherine S; Greenspan, Phillip; Pegg, Ronald B

    2016-12-01

    Inflammation is linked to numerous chronic disease states. Phenolic compounds have attracted attention because a number of these compounds possess anti-inflammatory properties. A phenolic crude extract was prepared from pecans and separated by Sephadex LH-20 column chromatography into low- and high-molecular-weight (LMW/HMW) fractions. Anti-inflammatory properties of these fractions were assessed in LPS-stimulated RAW 264.7 murine macrophage cells. NO and reactive oxygen species (ROS) production was monitored after 3 different experimental protocols: (1) pre-treatment with Escherichia coli O111:B4 lipopolysaccharide (LPS); (2) pre-treatment with a pecan crude extract and its fractions; and (3) co-incubation of LPS with a pecan crude extract and its fractions. The LMW fraction displayed a dose-dependent decrease in NO production and a significant decrease from the LPS control in ROS production when cells were either co-incubated with or pre-treated with LPS. The phenolics were characterized by HPLC to help identify those responsible for the observed effect. PMID:27374584

  5. Study on Association between Single Nucleotide Polymorphisms in Murine Double Minute 2 and Susceptibility of Hepatocellular Carcinoma

    Xia Wang

    2014-03-01

    Full Text Available Objective: To investigate the relationship between single nucleotide polymorphisms (SNP in murine double minute 2 (MDM2 and susceptibility and biological behavior of hepatocellularcarcinoma (HCC. Methods: MDM2 (rs2279744 site polymorphism in peripheral blood from 166 patients with HCC and 157 healthy controls were detected by SYBR GREEN PCR method and the relationship between MDM2 polymorphism and susceptibility and biological behavior of HCC was analyzed by comparing the differences of genotypes in two populations. Results: There was no statistical significance between two groups in terms of MDM2 allele distribution in research population (P = 0.753. The risk of HCC onset in individuals with GG+ TG genotype was 1.698 times of those with TT genotype in case group (95%CI = 1.027 -2.808. MDM2 SNP was associated with HBV infection and the degree of tumor differentiation (P< 0.05. The incidence of alleles in experimental group (T, 0.49; G, 0.51 was very different from that in control group (T, 0.59; G, 0.41 (P = 0.015. The incidence of GG genotype in patients with HCC (22.29% was significantly higher than those without HCC (13.38%. Compared with TT genotype, G allele or GG genotype had more correlation with HCC onset. Conclusion: Compared with TT genotype, MDM2 promoter SNP309 G allele or GG genotype is more associated with HCC onset in Chinese population.

  6. Immunoprophylactic potential of wheat grass extract on benzene-induced leukemia: An in vivo study on murine model

    Neelofar Khan

    2015-01-01

    Full Text Available Objectives: Wheat grass (Triticum aestivum is a gift of nature given to mankind. A number of scientific research on wheatgrass establishes its anticancer and antioxidant potential. Current work was focused to determine antileukemic effect of wheat grass. Materials and Methods: The commercial wheatgrass powder was extracted with 95% of methanol. Methanol extract of wheat grass was studied for acute oral toxicity as per revised Organization for Economic Cooperation and Development Guidelines number 423. Leukemia was successfully induced in Wister rats by intravenous injection of benzene. The blood was collected and analyzed for hematological parameters. Phagocytotic activity of the extract was determined. Results : Phytochemical screening revealed the presence of flavonoids, phenolics, carbohydrates, and amino acids. From acute toxicity studies, it was found that the methanol extract of wheatgrass was safe up to a dose level of 2000 mg/kg of body weight. Outcomes of hematological parameters in various experimental groups of murine model demonstrated antileukemic effect of extract. Methanol extract of wheatgrass aroused the process of phagocytosis of killed Candida albicans and also demonstrated a significant chemotactic activity at all tested concentrations. Conclusion: In the current work, methanol extract of wheat grass demonstrated antileukemic potential that might be due to the presence of flavonoids and polyphenolics in it. Further isolation, structural characterization of active constituents is necessary to extrapolate the mechanism of action.

  7. Systems genetics of liver fibrosis: identification of fibrogenic and expression quantitative trait loci in the BXD murine reference population.

    Rabea A Hall

    Full Text Available The progression of liver fibrosis in response to chronic injury varies considerably among individual patients. The underlying genetics is highly complex due to large numbers of potential genes, environmental factors and cell types involved. Here, we provide the first toxicogenomic analysis of liver fibrosis induced by carbon tetrachloride in the murine 'genetic reference panel' of recombinant inbred BXD lines. Our aim was to define the core of risk genes and gene interaction networks that control fibrosis progression. Liver fibrosis phenotypes and gene expression profiles were determined in 35 BXD lines. Quantitative trait locus (QTL analysis identified seven genomic loci influencing fibrosis phenotypes (pQTLs with genome-wide significance on chromosomes 4, 5, 7, 12, and 17. Stepwise refinement was based on expression QTL mapping with stringent selection criteria, reducing the number of 1,351 candidate genes located in the pQTLs to a final list of 11 cis-regulated genes. Our findings demonstrate that the BXD reference population represents a powerful experimental resource for shortlisting the genes within a regulatory network that determine the liver's vulnerability to chronic injury.

  8. Systems genetics of liver fibrosis: identification of fibrogenic and expression quantitative trait loci in the BXD murine reference population.

    Hall, Rabea A; Liebe, Roman; Hochrath, Katrin; Kazakov, Andrey; Alberts, Rudi; Laufs, Ulrich; Böhm, Michael; Fischer, Hans-Peter; Williams, Robert W; Schughart, Klaus; Weber, Susanne N; Lammert, Frank

    2014-01-01

    The progression of liver fibrosis in response to chronic injury varies considerably among individual patients. The underlying genetics is highly complex due to large numbers of potential genes, environmental factors and cell types involved. Here, we provide the first toxicogenomic analysis of liver fibrosis induced by carbon tetrachloride in the murine 'genetic reference panel' of recombinant inbred BXD lines. Our aim was to define the core of risk genes and gene interaction networks that control fibrosis progression. Liver fibrosis phenotypes and gene expression profiles were determined in 35 BXD lines. Quantitative trait locus (QTL) analysis identified seven genomic loci influencing fibrosis phenotypes (pQTLs) with genome-wide significance on chromosomes 4, 5, 7, 12, and 17. Stepwise refinement was based on expression QTL mapping with stringent selection criteria, reducing the number of 1,351 candidate genes located in the pQTLs to a final list of 11 cis-regulated genes. Our findings demonstrate that the BXD reference population represents a powerful experimental resource for shortlisting the genes within a regulatory network that determine the liver's vulnerability to chronic injury. PMID:24586654

  9. Chemokine binding protein M3 of murine gammaherpesvirus 68 modulates the host response to infection in a natural host.

    David J Hughes

    2011-03-01

    Full Text Available Murine γ-herpesvirus 68 (MHV-68 infection of Mus musculus-derived strains of mice is an attractive model of γ-herpesvirus infection. Surprisingly, however, ablation of expression of MHV-68 M3, a secreted protein with broad chemokine-binding properties in vitro, has no discernable effect during experimental infection via the respiratory tract. Here we demonstrate that M3 indeed contributes significantly to MHV-68 infection, but only in the context of a natural host, the wood mouse (Apodemus sylvaticus. Specifically, M3 was essential for two features unique to the wood mouse: virus-dependent inducible bronchus-associated lymphoid tissue (iBALT in the lung and highly organized secondary follicles in the spleen, both predominant sites of latency in these organs. Consequently, lack of M3 resulted in substantially reduced latency in the spleen and lung. In the absence of M3, splenic germinal centers appeared as previously described for MHV-68-infected laboratory strains of mice, further evidence that M3 is not fully functional in the established model host. Finally, analyses of M3's influence on chemokine and cytokine levels within the lungs of infected wood mice were consistent with the known chemokine-binding profile of M3, and revealed additional influences that provide further insight into its role in MHV-68 biology.

  10. Chemokine binding protein M3 of murine gammaherpesvirus 68 modulates the host response to infection in a natural host.

    Hughes, David J; Kipar, Anja; Leeming, Gail H; Bennett, Elaine; Howarth, Deborah; Cummerson, Joanne A; Papoula-Pereira, Rita; Flanagan, Brian F; Sample, Jeffery T; Stewart, James P

    2011-03-01

    Murine γ-herpesvirus 68 (MHV-68) infection of Mus musculus-derived strains of mice is an attractive model of γ-herpesvirus infection. Surprisingly, however, ablation of expression of MHV-68 M3, a secreted protein with broad chemokine-binding properties in vitro, has no discernable effect during experimental infection via the respiratory tract. Here we demonstrate that M3 indeed contributes significantly to MHV-68 infection, but only in the context of a natural host, the wood mouse (Apodemus sylvaticus). Specifically, M3 was essential for two features unique to the wood mouse: virus-dependent inducible bronchus-associated lymphoid tissue (iBALT) in the lung and highly organized secondary follicles in the spleen, both predominant sites of latency in these organs. Consequently, lack of M3 resulted in substantially reduced latency in the spleen and lung. In the absence of M3, splenic germinal centers appeared as previously described for MHV-68-infected laboratory strains of mice, further evidence that M3 is not fully functional in the established model host. Finally, analyses of M3's influence on chemokine and cytokine levels within the lungs of infected wood mice were consistent with the known chemokine-binding profile of M3, and revealed additional influences that provide further insight into its role in MHV-68 biology. PMID:21445235

  11. THE IMMUNE RESPONSE TO PARASITIC HELMINTHS: INSIGHT FROM MURINE MODELS.

    Helminth parasites are a large group of multi-cellular organisms that affect vast numbers of humans and are a major cause of disease. Several relevant experimental mouse models, representing the spectrum of human disease, have been very helpful in analyzing and characterizing the host immune respon...

  12. Comparative effects in vivo of recombinant murine interleukin 3, natural murine colony-stimulating factor-1, and recombinant murine granulocyte-macrophage colony-stimulating factor on myelopoiesis in mice.

    Broxmeyer, H E; Williams, D.E.; Cooper, S; Shadduck, R K; Gillis, S.; Waheed, A.; Urdal, D L; Bicknell, D C

    1987-01-01

    Purified murine colony-stimulating factors (CSF) recombinant interleukin 3 (IL-3), natural CSF-1, and recombinant granulocyte-macrophage (GM) CSF were assessed in vivo for their effects on BDF1 mouse bone marrow and spleen granulocyte-macrophage (CFU-GM), erythroid (BFU-E), and multipotential (CFU-GEMM) progenitor cells in untreated mice and in mice pretreated with purified iron-saturated human lactoferrin (LF). The CSF and LF preparations did not contain detectable endotoxin (less than 0.1 n...

  13. Soluble interleukin 2 receptors are released from the cell surface of normal murine B lymphocytes stimulated with interleukin 5.

    Loughnan, M S; Sanderson, C. J.; Nossal, G J

    1988-01-01

    Murine T and B lymphocytes can be induced to release soluble interleukin 2 receptors (IL2Rs). This receptor is believed to be a truncated form of the 55-kDa chain of the cell-membrane-associated receptor. It has been speculated that this receptor may play an important immunoregulatory role by binding to interleukin 2 (IL-2). We report here that interleukin 5 can induce normal murine B cells to release soluble IL2Rs. This extends our finding that interleukin 5 similarly can induce murine B cel...

  14. Inflammatory manifestations of experimental lymphatic insufficiency.

    Raymond Tabibiazar

    2006-07-01

    Full Text Available BACKGROUND: Sustained lymph stagnation engenders a pathological response that is complex and not well characterized. Tissue inflammation in lymphedema may reflect either an active or passive consequence of impaired immune traffic. METHODS AND FINDINGS: We studied an experimental model of acute post-surgical lymphedema in the tails of female hairless, immunocompetent SKH-1 mice. We performed in vivo imaging of impaired immune traffic in experimental, murine acquired lymphatic insufficiency. We demonstrated impaired mobilization of immunocompetent cells from the lymphedematous region. These findings correlated with histopathological alterations and large-scale transcriptional profiling results. We found intense inflammatory changes in the dermis and the subdermis. The molecular pattern in the RNA extracted from the whole tissue was dominated by the upregulation of genes related to acute inflammation, immune response, complement activation, wound healing, fibrosis, and oxidative stress response. CONCLUSIONS: We have characterized a mouse model of acute, acquired lymphedema using in vivo functional imaging and histopathological correlation. The model closely simulates the volume response, histopathology, and lymphoscintigraphic characteristics of human acquired lymphedema, and the response is accompanied by an increase in the number and size of microlymphatic structures in the lymphedematous cutaneous tissues. Molecular characterization through clustering of genes with known functions provides insights into processes and signaling pathways that compose the acute tissue response to lymph stagnation. Further study of genes identified through this effort will continue to elucidate the molecular mechanisms and lead to potential therapeutic strategies for lymphatic vascular insufficiency.

  15. Dual effects of vitamin D-induced alteration of TH1/TH2 cytokine expression: enhancing IgE production and decreasing airway eosinophilia in murine allergic airway disease

    Matheu, Victor; Bäck, Ove; Mondoc, Emma;

    2003-01-01

    BACKGROUND: Vitamin D, a common food additive, has been shown to prevent the induction of experimental autoimmune diseases in mice. A possible immune deviation from T(H)1 to T(H)2 responses has been postulated. Although there is no doubt about the beneficial effects of vitamin D, its role in...... allergy has not been investigated. OBJECTIVE: To define the role of vitamin D in modulating the development of a T(H)2-mediated disease, we used a murine model of pulmonary eosinophilic inflammation. METHODS: Five-week-old mice were primed on day 0 with ovalbumin intraperitoneally. Then they were nasally...... hold promising beneficial effects in airway eosinophilia....

  16. Rosiglitazone inhibits metastasis development of a murine mammary tumor cell line LMM3

    Rolando Romina

    2008-02-01

    Full Text Available Abstract Background Activation of peroxisome proliferator-activated receptors γ (PPARγ induces diverse effects on cancer cells. The thiazolidinediones (TZDs, such as troglitazone and ciglitazone, are PPARγ agonists exhibiting antitumor activities; however, the underlying mechanism remains inconclusive. Rosiglitazone (RGZ, a synthetic ligand of PPARγ used in the treatment of Type 2 diabetes, inhibits growth of some tumor cells and is involved in other processes related to cancer progression. Opposing results have also been reported with different ligands on tumor cells. The purpose of this study was to determine if RGZ and 15d-PGJ2 induce antitumor effects in vivo and in vitro on the murine mammary tumor cell line LMM3. Methods The effect on LMM3 cell viability and nitric oxide (NO production of different doses of RGZ, 15-dPGJ2, BADGE and GW9662 were determined using the MTS colorimetric assay and the Griess reaction respectively. In vivo effect of orally administration of RGZ on tumor progression was evaluated either on s.c. primary tumors as well as on experimental metastasis. Cell adhesion, migration (wound assay and invasion in Transwells were performed. Metalloproteinase activity (MMP was determined by zymography in conditioned media from RGZ treated tumor cells. PPARγ expression was detected by inmunohistochemistry in formalin fixed tumors and by western blot in tumor cell lysates. Results RGZ orally administered to tumor-bearing mice decreased the number of experimental lung metastases without affecting primary s.c. tumor growth. Tumor cell adhesion and migration, as well as metalloproteinase MMP-9 activity, decreased in the presence of 1 μM RGZ (non-cytotoxic dose. RGZ induced PPARγ protein expression in LMM3 tumors. Although metabolic activity -measured by MTS assay- diminished with 1–100 μM RGZ, 1 μM-treated cells recovered their proliferating capacity while 100 μM treated cells died. The PPARγ antagonist Biphenol A

  17. Rosiglitazone inhibits metastasis development of a murine mammary tumor cell line LMM3

    Activation of peroxisome proliferator-activated receptors γ (PPARγ) induces diverse effects on cancer cells. The thiazolidinediones (TZDs), such as troglitazone and ciglitazone, are PPARγ agonists exhibiting antitumor activities; however, the underlying mechanism remains inconclusive. Rosiglitazone (RGZ), a synthetic ligand of PPARγ used in the treatment of Type 2 diabetes, inhibits growth of some tumor cells and is involved in other processes related to cancer progression. Opposing results have also been reported with different ligands on tumor cells. The purpose of this study was to determine if RGZ and 15d-PGJ2 induce antitumor effects in vivo and in vitro on the murine mammary tumor cell line LMM3. The effect on LMM3 cell viability and nitric oxide (NO) production of different doses of RGZ, 15-dPGJ2, BADGE and GW9662 were determined using the MTS colorimetric assay and the Griess reaction respectively. In vivo effect of orally administration of RGZ on tumor progression was evaluated either on s.c. primary tumors as well as on experimental metastasis. Cell adhesion, migration (wound assay) and invasion in Transwells were performed. Metalloproteinase activity (MMP) was determined by zymography in conditioned media from RGZ treated tumor cells. PPARγ expression was detected by inmunohistochemistry in formalin fixed tumors and by western blot in tumor cell lysates. RGZ orally administered to tumor-bearing mice decreased the number of experimental lung metastases without affecting primary s.c. tumor growth. Tumor cell adhesion and migration, as well as metalloproteinase MMP-9 activity, decreased in the presence of 1 μM RGZ (non-cytotoxic dose). RGZ induced PPARγ protein expression in LMM3 tumors. Although metabolic activity -measured by MTS assay- diminished with 1–100 μM RGZ, 1 μM-treated cells recovered their proliferating capacity while 100 μM treated cells died. The PPARγ antagonist Biphenol A diglicydyl ether (BADGE) did not affect RGZ activity. On

  18. The Relationship between the Antitumor Effect of the IL-12 Gene Therapy and the Expression of Th1 Cytokines in an HPV16-Positive Murine Tumor Model

    Flor García Paz

    2014-01-01

    Full Text Available Objective. The goal of the present study was to investigate the effect of IL-12 expressed in plasmid on the Th1 cytokine profile in an experimental HPV16-positive murine tumor model and the association with the IL-12’s antitumor effect. Methods. Mice were injected with BMK-16/myc cells to establish HPV16-positive tumor and then pNGVL3-mIL-12 plasmid; pcDNA3 plasmid or PBS was injected directly into tumor site. The antitumor effect of the treatment was evaluated and the cytokines expression profile in each tumor tissue was analyzed. Results. Treatment with pNGVL3-mIL-12 plasmid had a significant antitumor effect, and a Th2-Th3-type cytokines prolife was detected in the murine tumor model with expression of the cytokines IL-10, IL-4, and TGF-β1. However, after the tumor was treated with three intratumoral injections of plasmid containing IL-12 cDNA, it showed a cytokine profile associated with Th1 with expression of IL-2, IL-12, and IFN-γ cytokines and reduced expression of IL-10, IL-4, and TGF-β1. Conclusions. The treatment with the IL-12 gene in the experimental HPV16-positive tumor model promoted the activation of the cellular immune response via expression of a Th1-type cytokine profile and was associated with the inhibition of tumor growth. Thus, IL-12 treatment represents a novel approach for gene therapy against cervical cancer.

  19. Peroxisome Proliferator-activated Receptor-γ Coactivator 1-α (PGC1α) Protects against Experimental Murine Colitis.

    Cunningham, Kellie E; Vincent, Garret; Sodhi, Chhinder P; Novak, Elizabeth A; Ranganathan, Sarangarajan; Egan, Charlotte E; Stolz, Donna Beer; Rogers, Matthew B; Firek, Brian; Morowitz, Michael J; Gittes, George K; Zuckerbraun, Brian S; Hackam, David J; Mollen, Kevin P

    2016-05-01

    Peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC1α) is the primary regulator of mitochondrial biogenesis and was recently found to be highly expressed within the intestinal epithelium. PGC1α is decreased in the intestinal epithelium of patients with inflammatory bowel disease, but its role in pathogenesis is uncertain. We now hypothesize that PGC1α protects against the development of colitis and helps to maintain the integrity of the intestinal barrier. We selectively deleted PGC1α from the intestinal epithelium of mice by breeding a PGC1α(loxP/loxP) mouse with a villin-cre mouse. Their progeny (PGC1α(ΔIEC) mice) were subjected to 2% dextran sodium sulfate (DSS) colitis for 7 days. The SIRT1 agonist SRT1720 was used to enhance PGC1α activation in wild-type mice during DSS exposure. Mice lacking PGC1α within the intestinal epithelium were more susceptible to DSS colitis than their wild-type littermates. Pharmacologic activation of PGC1α successfully ameliorated disease and restored mitochondrial integrity. These findings suggest that a depletion of PGC1α in the intestinal epithelium contributes to inflammatory changes through a failure of mitochondrial structure and function as well as a breakdown of the intestinal barrier, which leads to increased bacterial translocation. PGC1α induction helps to maintain mitochondrial integrity, enhance intestinal barrier function, and decrease inflammation. PMID:26969166

  20. Utility of the Microculture Method in Non-Invasive Samples Obtained from an Experimental Murine Model with Asymptomatic Leishmaniasis

    Allahverdiyev, Adil M; Bagirova, Malahat; Cakir-Koc, Rabia; Elcicek, Serhat; Oztel, Olga Nehir; Canim-Ates, Sezen; Abamor, Emrah Sefik; Yesilkir-Baydar, Serap

    2012-01-01

    The sensitivity of diagnostic methods for visceral leishmaniasis (VL) decreases because of the low number of parasites and antibody amounts in asymptomatic healthy donors who are not suitable for invasive sample acquisition procedures. Therefore, new studies are urgently needed to improve the sensitivity and specificity of the diagnostic approaches in non-invasive samples. In this study, the sensitivity of the microculture method (MCM) was compared with polymerase chain reaction (PCR), enzyme...

  1. Kinetics of expression of costimulatory molecules and their ligands in murine relapsing experimental autoimmune encephalomyelitis in vivo

    Issazadeh-Navikas, Shohreh; Navikas, V; Schaub, M; Sayegh, M; Khoury, S

    1998-01-01

    -2, had distinct expression patterns in the CNS; CD28 was highly expressed and correlated with B7-2 expression on APCs (macrophages/microglia as well as astrocytes) and with the clinical signs of EAE. CTLA4, on the other hand, was expressed by substantially fewer cells during the effector phase of disease...... autoimmune disease model characterized by remissions and relapses. Our data suggest that the targeting of costimulatory molecules to block an immune response must take into account the expression patterns in the target organ....

  2. Oral administration of Parabacteroides distasonis antigens attenuates experimental murine colitis through modulation of immunity and microbiota composition

    Kverka, Miloslav; Zákostelská, Zuzana; Klimešová, Klára; Sokol, Dan; Hudcovic, Tomáš; Hrnčíř, Tomáš; Rossmann, Pavel; Mrázek, Jakub; Kopečný, Jan; Verdu, E.; Tlaskalová-Hogenová, Helena

    2011-01-01

    Roč. 163, č. 2 (2011), s. 250-259. ISSN 0009-9104 R&D Projects: GA AV ČR KJB500200904; GA AV ČR 1QS500200572; GA AV ČR IAA5020205; GA ČR GD310/08/H077; GA ČR GA303/08/0367; GA ČR GA305/08/0535 Institutional research plan: CEZ:AV0Z50200510; CEZ:AV0Z50450515 Keywords : animal models * cytokines/interleukins * DSS colitis Subject RIV: EE - Microbiology, Virology Impact factor: 3.360, year: 2011

  3. The effect of vitamin A supplementation and diphtheria-tetanus-pertussis vaccination on parasitaemia in an experimental murine malaria model

    Jørgensen, Mathias Jul; Hein-Kristensen, Line; Hempel, Casper;

    2011-01-01

    infectious diseases when given with the diphtheria-tetanus-pertussis (DTP) vaccine. The immunological effects of combining the 2 treatments are unknown. Methods: We studied the effect of treating C57BL/6 mice with VAS and DTP, 1 week prior to infection with Plasmodium berghei ANKA. The progression of disease...

  4. Efficacies of KY62 against Leishmania amazonensis and Leishmania donovani in Experimental Murine Cutaneous Leishmaniasis and Visceral Leishmaniasis

    Al-Abdely, Hail M.; Graybill, John R.; Bocanegra, Rosie; Najvar, Laura; Montalbo, Eleanor; Regen, Steven L.; Melby, Peter C.

    1998-01-01

    Current therapy for leishmaniasis is unsatisfactory because parenteral antimonial salts and pentamidine are associated with significant toxicity and failure rates. We examined the efficacy of KY62, a new, water-soluble, polyene antifungal, against cutaneous infection with Leishmania amazonensis and against visceral infection with Leishmania donovani in susceptible BALB/c mice. Mice were infected with L. amazonensis promastigotes in the ear pinna and in the tail and were treated with KY62 or a...

  5. BENSC experimental reports 1993

    This volume contains the guest groups' experimental reports describing experimental work carried out on the Berlin Scattering Center in 1993. These experimental reports are intended as interim summaries. (HP)

  6. BENSC. Experimental reports 1994

    This volume contains the guest groups' experimental reports describing experimental work carried out on the Berlin Scattering Center in 1994. These experimental reports are intended as interim summaries. (HP)

  7. Structural and functional characterization of human and murine C5a anaphylatoxins

    Schatz-Jakobsen, Janus Asbjørn; Yatime, Laure, E-mail: lay@mb.au.dk; Larsen, Casper [Aarhus University, Gustav Wieds Vej 10C, DK-8000 Aarhus (Denmark); Petersen, Steen Vang [Aarhus University, Bartholin Building, Wilhelm Meyers Allé 4, DK-8000 Aarhus (Denmark); Klos, Andreas [Medical School Hannover, Hannover (Germany); Andersen, Gregers Rom, E-mail: lay@mb.au.dk [Aarhus University, Gustav Wieds Vej 10C, DK-8000 Aarhus (Denmark)

    2014-06-01

    The structure of the human C5aR antagonist, C5a-A8, reveals a three-helix bundle conformation similar to that observed for human C5a-desArg, whereas murine C5a and C5a-desArg both form the canonical four-helix bundle. These conformational differences are discussed in light of the differential C5aR activation properties observed for the human and murine complement anaphylatoxins across species. Complement is an ancient part of the innate immune system that plays a pivotal role in protection against invading pathogens and helps to clear apoptotic and necrotic cells. Upon complement activation, a cascade of proteolytic events generates the complement effectors, including the anaphylatoxins C3a and C5a. Signalling through their cognate G-protein coupled receptors, C3aR and C5aR, leads to a wide range of biological events promoting inflammation at the site of complement activation. The function of anaphylatoxins is regulated by circulating carboxypeptidases that remove their C-terminal arginine residue, yielding C3a-desArg and C5a-desArg. Whereas human C3a and C3a-desArg adopt a canonical four-helix bundle fold, the conformation of human C5a-desArg has recently been described as a three-helix bundle. Here, the crystal structures of an antagonist version of human C5a, A8{sup Δ71–73}, and of murine C5a and C5a-desArg are reported. Whereas A8{sup Δ71–73} adopts a three-helix bundle conformation similar to human C5a-desArg, the two murine proteins form a four-helix bundle. A cell-based functional assay reveals that murine C5a-desArg, in contrast to its human counterpart, exerts the same level of activition as murine C5a on its cognate receptor. The role of the different C5a conformations is discussed in relation to the differential activation of C5a receptors across species.

  8. EFFECTS OF IMMUNOSUPPRESSION WITH CYCLOPHOSPHAMIDE ON ACUTE MURINE CYTOMEGALOVIRUS INFECTION AND VIRUS-AUGMENTED NATURAL KILLER CELL ACTIVITY

    The effects of cyclophosphamide (CY) treatment on acute murine cytomegalovirus (MCMV) infection were studied to explore the potential usefulness of MCMV as a means of detecting immune dysfunction and to identify host defense mechanisms important for protection against MCMV.

  9. Effect of Control-released Basic Fibroblast Growth Factor Incorporated in β-Tricalcium Phosphate for Murine Cranial Model

    Azusa Shimizu, MD

    2014-03-01

    Conclusions: These findings suggest that control-released bFGF incorporated in β-TCP can accelerate bone regeneration in the murine cranial defect model and may be promising for the clinical treatment of cranial defects.

  10. Efficacy of paracetamol on patent ductus arteriosus closure may be dose dependent: evidence from human and murine studies.

    El-Khuffash, Afif

    2014-09-01

    We evaluated the clinical effectiveness of variable courses of paracetamol on patent ductus arteriosus (PDA) closure and examined its effect on the in vitro term and preterm murine ductus arteriosus (DA).

  11. 3-D QSAR ANALYSIS OF INHIBITION OF MURINE SOLUBLE EPOXIDE HYDROLASE (MSEH) BY BENZOYLUREAS, ARYLUREAS, AND THEIR ANALOGUES. (R825433)

    Two hundred and seventy-one compounds including benzoylureas, arylureas and related compounds were assayed using recombinant murine soluble epoxide hydrolase (MsEH) produced from a baculovirus expression system. Among all the insect growth regulators assayed, 18 benzoylphenylu...

  12. Detection of adulterated murine components in meat products by TaqMan© real-time PCR.

    Fang, Xin; Zhang, Chi

    2016-02-01

    Using murine meat to substitute mutton has been identified as a new type of meat fraud in China, yet no detection method for murine species has been reported. Here, three kinds of rodent were used as target species to establish a murine-specific real-time PCR method of detection. The mitochondrial cytochrome b gene (cytb) of each target was sequenced and a TaqMan probe was designed based on the cytb. Simultaneously, an internal positive control (IPC) plasmid along with its respective probe were designed to monitor the PCR reaction. As a result, the duplex real-time PCR system was verified to be specific. The limit of detection (LOD) was lower than 1 pg of DNA per reaction and 0.1% murine contamination in meat mixtures. Standard curves were generated for a quantitative analysis. Thus, this study provided a new tool to control the quality of meat products for official and third-party laboratories. PMID:26304376

  13. Eicosapentaenoic and docosahexaenoic acid ethyl esters differentially enhance B-cell activity in murine obesity[S

    Teague, Heather; Harris, Mitchel; Fenton, Jenifer; Lallemand, Perrine; Shewchuk, Brian M.; Shaikh, Saame Raza

    2014-01-01

    EPA and DHA are not biologically equivalent; however, their individual activity on B cells is unknown. We previously reported fish oil enhanced murine B-cell activity in obesity. To distinguish between the effects of EPA and DHA, we studied the ethyl esters of EPA and DHA on murine B-cell function as a function of time. We first demonstrate that EPA and DHA maintained the obese phenotype, with no improvements in fat mass, adipose inflammatory cytokines, fasting insulin, or glucose clearance. ...

  14. Murine patellar tendon biomechanical properties and regional strain patterns during natural tendon-to-bone healing after acute injury

    Gilday, Steven D.; Casstevens, E. Chris; Kenter, Keith; Jason T Shearn; David L Butler

    2013-01-01

    Tendon-to-bone healing following acute injury is generally poor and often fails to restore normal tendon biomechanical properties. In recent years, the murine patellar tendon (PT) has become an important model system for studying tendon healing and repair due to its genetic tractability and accessible location within the knee. However, the mechanical properties of native murine PT, specifically the regional differences in tissue strains during loading, and the biomechanical outcomes of natura...

  15. Visualization of Intrapulmonary Lymph Vessels in Healthy and Inflamed Murine Lung Using CD90/Thy-1 as a Marker

    Kretschmer, Sarah; Dethlefsen, Ina; Hagner-Benes, Stefanie; Marsh, Leigh M; Garn, Holger; König, Peter

    2013-01-01

    Background Lymphatic vessels play a pivotal role in fluid drainage and egress of immune cells from the lung. However, examining murine lung lymphatics is hampered by the expression of classical lymph endothelial markers on other cell types, which hinders the unambiguous identification of lymphatics. The expression of CD90/Thy-1 on lymph endothelium was recently described and we therefore examined its suitability to identify murine pulmonary lymph vessels under healthy and inflammatory conditi...

  16. Retrovirus-mediated gene transfer of human adenosine deaminase: expression of functional enzyme in murine hematopoietic stem cells in vivo.

    Lim, B; Williams, D A; Orkin, S H

    1987-01-01

    Simplified Moloney murine leukemia virus-based recombinant retrovirus vectors have been constructed which transduce human adenosine deaminase (ADA) cDNA. ADA transcription is under the control of the constitutive promoter for the human X chromosome phosphoglycerate kinase (pgk) gene. In these simplified vectors, dominant selectable markers are not included and selection is dependent on overproduction of functional ADA enzyme. Primary murine hematopoietic cells were infected with helper-free r...

  17. Inhibition of Src kinase activity attenuates amyloid associated microgliosis in a murine model of Alzheimer’s disease

    Dhawan Gunjan; Combs Colin K

    2012-01-01

    Abstract Background Microglial activation is an important histologic characteristic of the pathology of Alzheimer’s disease (AD). One hypothesis is that amyloid beta (Aβ) peptide serves as a specific stimulus for tyrosine kinase-based microglial activation leading to pro-inflammatory changes that contribute to disease. Therefore, inhibiting Aβ stimulation of microglia may prove to be an important therapeutic strategy for AD. Methods Primary murine microglia cultures and the murine microglia c...

  18. Retrovirus vectors containing an internal attachment site: evidence that circles are not intermediates to murine retrovirus integration.

    Ellis, J.; Bernstein, A.

    1989-01-01

    Murine cells were infected with a retrovirus vector containing a defective native attachment (att) site, an internal att site, and a neo gene. Analysis of the proviruses by virus rescue and Southern blots demonstrated that internal att sites were not utilized for integration and could not complement defects in the native site. These data suggest that murine retroviruses do not integrate in vivo through tandem long terminal repeat circular DNA intermediates.

  19. Selective host range restriction of goat cells for recombinant murine leukemia virus and feline leukemia virus type A.

    Fischinger, P J; Thiel, H J; Blevins, C S; Dunlop, N M

    1981-01-01

    We isolated a strain of normal goat fibroblasts which was uniquely selective in that it allowed the replication of xenotropic murine leukemia virus but not polytropic recombinant murine leukemia virus. In addition, feline leukemia virus type A replication was severely diminished in these goat cells, whereas feline leukemia virus type B and feline endogenous RD114-CCC viruses replicated efficiently. No other known cells exhibit this pattern of virus growth restriction. These goat cells allow t...

  20. Probiotic yeasts: Anti-inflammatory potential of various non-pathogenic strains in experimental colitis in mice

    Benot; Foligné; Jo■lle; Dewulf; Pascal; Vandekerckove; Georges; Pignède; Bruno; Pot

    2010-01-01

    AIM: To evaluate the in vitro immunomodulation capacity of various non-pathogenic yeast strains and to investigate the ability of some of these food grade yeasts to prevent experimental colitis in mice.METHODS: In vitro immunomodulation was assessed by measuring cytokines [interleukin (IL)-12p70,IL-10,tumor necrosis factor and interferon γ] released by human peripheral blood mononuclear cells after 24 h stimulation with 6 live yeast strains (Saccharomyces ssp.) and with bacterial reference strains.A murine ...

  1. Comparison of the chromosomal localization of murine and human glucocerebrosidase genes and of the deduced amino acid sequences

    To study structure-function relationships and molecular evolution, the authors determined the nucleotide sequence and chromosomal location of the gene encoding murine glucocerebrosidase. In the protein coding region of the murine cDNA, the nucleotide sequence and the corresponding deduced amino acid sequences were 82% and 86% identical to the respective humans sequences. All five amino acids presently known to be essential for normal enzymatic activity were conserved between mouse and man. The murine enzyme had a single deletion relative to the human enzyme at amino acid number 273. One ATG translation initiation signal was present in the mouse sequence in contrast to the human sequence, where two start codons have been reported. Nucleotide sequencing of a clone derived from murine genomic DNA revealed that the murine signal for translation initiation was located in exon 2. The locations of all 10 introns were conserved among mouse and man. They mapped the genetic locus for glucocerebrosidase to mouse chromosome 3, at a position 7.6 ± 3.2 centimorgans from the locus for the β subunit of nerve growth factor. Comparison of linkage relationships in the human and murine genome indicates that these closely linked mouse genes are also syntenic on human chromosome 1 but in positions that span the centromere

  2. Murine Hind Limb Long Bone Dissection and Bone Marrow Isolation.

    Amend, Sarah R; Valkenburg, Kenneth C; Pienta, Kenneth J

    2016-01-01

    Investigation of the bone and the bone marrow is critical in many research fields including basic bone biology, immunology, hematology, cancer metastasis, biomechanics, and stem cell biology. Despite the importance of the bone in healthy and pathologic states, however, it is a largely under-researched organ due to lack of specialized knowledge of bone dissection and bone marrow isolation. Mice are a common model organism to study effects on bone and bone marrow, necessitating a standardized and efficient method for long bone dissection and bone marrow isolation for processing of large experimental cohorts. We describe a straightforward dissection procedure for the removal of the femur and tibia that is suitable for downstream applications, including but not limited to histomorphologic analysis and strength testing. In addition, we outline a rapid procedure for isolation of bone marrow from the long bones via centrifugation with limited handling time, ideal for cell sorting, primary cell culture, or DNA, RNA, and protein extraction. The protocol is streamlined for rapid processing of samples to limit experimental error, and is standardized to minimize user-to-user variability. PMID:27168390

  3. Cáncer experimental e inflamación sistémica en un modelo murino Systemic inflammation and experimental cancer in a murine model

    Juan Bruzzo; Paula Chiarella; Gabriela Fernández; Oscar D. Bustuoabad; Raúl A. Ruggiero

    2007-01-01

    La asociación entre cáncer e inflamación en un órgano o tejido se encuentra sólidamente establecida. En efecto, se sabe que en sitios de inflamación crónica, existe una mayor probabilidad de que se origine un tumor y que procesos inflamatorios locales pueden acelerar el crecimiento de tumores preexistentes en animales y seres humanos. Por otro lado, la relación entre cáncer e inflamación sistémica ha sido menos estudiada. En este trabajo, demostramos que el crecimiento de un fibrosarcoma de r...

  4. Hematein, a casein kinase II inhibitor, inhibits lung cancer tumor growth in a murine xenograft model.

    Hung, Ming-Szu; Xu, Zhidong; Chen, Yu; Smith, Emmanuel; Mao, Jian-Hua; Hsieh, David; Lin, Yu-Ching; Yang, Cheng-Ta; Jablons, David M; You, Liang

    2013-11-01

    Casein kinase II (CK2) inhibitors suppress cancer cell growth. In this study, we examined the inhibitory effects of a novel CK2 inhibitor, hematein, on tumor growth in a murine xenograft model. We found that in lung cancer cells, hematein inhibited cancer cell growth, Akt/PKB Ser129 phosphorylation, the Wnt/TCF pathway and increased apoptosis. In a murine xenograft model of lung cancer, hematein inhibited tumor growth without significant toxicity to the mice tested. Molecular docking showed that hematein binds to CK2α in durable binding sites. Collectively, our results suggest that hematein is an allosteric inhibitor of protein kinase CK2 and has antitumor activity to lung cancer. PMID:24008396

  5. Differential regulation of follicle stimulating hormone by activin A and TGFB1 in murine gonadotropes

    Miller William L

    2005-12-01

    Full Text Available Abstract Background Activins stimulate the synthesis of follicle stimulating hormone (FSH in pituitary gonadotropes, at least in part, by inducing transcription of its beta subunit (Fshb. Evidence from several laboratories studying transformed murine LbetaT2 gonadotropes indicates that activins signal through Smad-dependent and/or Smad-independent pathways, similar to those used by transforming growth factor beta-1 (TGFB1 in other cell types. Therefore, given common intracellular signaling mechanisms of these two ligands, we examined whether TGFBs can also induce transcription of Fshb in LbetaT2 cells as well as in purified primary murine gonadotropes. Methods Murine Fshb promoter-reporter (-1990/+1 mFshb-luc activity was measured in LbetaT2 cells treated with activin A or TGFB1, and in cells transfected with either activin or TGFB receptors. The ability of the ligands to stimulate phosphorylation of Smads 2 and 3 in LbetaT2 cells was measured by western blot analysis, and expression of TGFB type I and II receptors was assessed by reverse transcriptase polymerase chain reaction in both LbetaT2 cells and primary gonadotropes purified from male mice of different ages. Finally, regulation of endogenous murine Fshb mRNA levels by activin A and TGFB1 in purified gonadotropes and whole pituitary cultures was measured using quantitative RT-PCR. Results Activin A dose-dependently stimulated -1990/+1 mFshb-luc activity in LbetaT2 cells, but TGFB1 had no effect at doses up to 5 nM. Similarly, activin A, but not TGFB1, stimulated Smad 2 and 3 phosphorylation in these cells. Constitutively active forms of the activin (Acvr1b-T206D and TGFB (TGFBR1-T204D type I receptors strongly stimulated -1990/+1 mFshb-luc activity, showing that mechanisms down stream of Tgfbr1 seem to be intact in LbetaT2 cells. RT-PCR analysis of LbetaT2 cells and whole adult murine pituitaries indicated that both expressed Tgfbr1 mRNA, but that Tgfbr2 was not detected in LbetaT2 cells

  6. Murine Mueller cells are progenitor cells for neuronal cells and fibrous tissue cells

    Mammalian Mueller cells have been reported to possess retinal progenitor cell properties and generate new neurons after injury. This study investigates murine Mueller cells under in vitro conditions for their capability of dedifferentiation into retinal progenitor cells. Mueller cells were isolated from mouse retina, and proliferating cells were expanded in serum-containing medium. For dedifferentiation, the cultured cells were transferred to serum-replacement medium (SRM) at different points in time after their isolation. Interestingly, early cell passages produced fibrous tissue in which extracellular matrix proteins and connective tissue markers were differentially expressed. In contrast, aged Mueller cell cultures formed neurospheres in SRM that are characteristic for neuronal progenitor cells. These neurospheres differentiated into neuron-like cells after cultivation on laminin/ornithine cell culture substrate. Here, we report for the first time that murine Mueller cells can be progenitors for both, fibrous tissue cells and neuronal cells, depending on the age of the cell culture

  7. Mutated primer binding sites interacting with different tRNAs allow efficient murine leukemia virus replication

    Lund, Anders Henrik; Duch, M; Lovmand, J;

    1993-01-01

    Two Akv murine leukemia virus-based retroviral vectors with primer binding sites matching tRNA(Gln-1) and tRNA(Lys-3) were constructed. The transduction efficiency of these mutated vectors was found to be comparable to that of a vector carrying the wild-type primer binding site matching t......RNA(Pro). Polymerase chain reaction amplification and sequence analysis of transduced proviruses confirmed the transfer of vectors with mutated primer binding sites and further showed that tRNA(Gln-2) may act efficiently in conjunction with the tRNA(Gln-1) primer binding site. We conclude that murine leukemia virus...... can replicate by using various tRNA molecules as primers and propose primer binding site-tRNA primer interactions to be of major importance for tRNA primer selection. However, efficient primer selection does not require perfect Watson-Crick base pairing at all 18 positions of the primer binding site....

  8. Eukaryotic expression, purification, crystallization and preliminary X-ray analysis of murine Manic Fringe

    Jinek, Martin [European Molecular Biology Laboratory (EMBL), Meyerhofstrasse 1, D-69117 Heidelberg (Germany); Conti, Elena, E-mail: conti@embl.de [European Molecular Biology Laboratory (EMBL), Meyerhofstrasse 1, D-69117 Heidelberg (Germany); Max Planck Institute of Biochemistry, Am Klopferspitz 18, D-82152 Martinsried (Germany)

    2006-08-01

    The catalytic domain of the murine glycosyltransferase Manic Fringe was expressed in insect cells. Removal by site-directed mutagenesis of two N-glycosylation sites present in the protein was essential to obtain crystals that diffracted to 1.8 Å resolution. Fringe proteins are Golgi-resident β1,3-N-acetylglucosaminyltransferases that regulate development in metazoa through glycosylation of the Notch receptor and its ligands. The catalytic domain of murine Manic Fringe was expressed in the baculovirus/insect-cell system as a secreted protein. Mass-spectrometric analysis of the purified protein indicated the presence of two N-linked glycans. Abolishing the glycosylation sites by site-directed mutagenesis was necessary in order to obtain orthorhombic crystals that diffracted to 1.8 Å resolution. For phasing, a highly redundant data set was collected using a crystal soaked with halide salts.

  9. Eukaryotic expression, purification, crystallization and preliminary X-ray analysis of murine Manic Fringe

    The catalytic domain of the murine glycosyltransferase Manic Fringe was expressed in insect cells. Removal by site-directed mutagenesis of two N-glycosylation sites present in the protein was essential to obtain crystals that diffracted to 1.8 Å resolution. Fringe proteins are Golgi-resident β1,3-N-acetylglucosaminyltransferases that regulate development in metazoa through glycosylation of the Notch receptor and its ligands. The catalytic domain of murine Manic Fringe was expressed in the baculovirus/insect-cell system as a secreted protein. Mass-spectrometric analysis of the purified protein indicated the presence of two N-linked glycans. Abolishing the glycosylation sites by site-directed mutagenesis was necessary in order to obtain orthorhombic crystals that diffracted to 1.8 Å resolution. For phasing, a highly redundant data set was collected using a crystal soaked with halide salts

  10. Developmentally-Dynamic Murine Brain Proteomes and Phosphoproteomes Revealed by Quantitative Proteomics

    Peter F. Doubleday

    2014-04-01

    Full Text Available Developmental processes are governed by a diverse suite of signaling pathways employing reversible phosphorylation. Recent advances in large-scale phosphoproteomic methodologies have made possible the identification and quantification of hundreds to thousands of phosphorylation sites from primary tissues. Towards a global characterization of proteomic changes across brain development, we present the results of a large-scale quantitative mass spectrometry study comparing embryonic, newborn and adult murine brain. Using anti-phosphotyrosine immuno-affinity chromatography and strong cation exchange (SCX chromatography, coupled to immobilized metal affinity chromatography (IMAC, we identified and quantified over 1,750 phosphorylation sites and over 1,300 proteins between three developmental states. Bioinformatic analyses highlight functions associated with the identified proteins and phosphoproteins and their enrichment at distinct developmental stages. These results serve as a primary reference resource and reveal dynamic developmental profiles of proteins and phosphoproteins from the developing murine brain.

  11. Different expression patterns of TRP genes in murine B and T lymphocytes

    A prolonged increase in the intracellular calcium concentration ([Ca2+]i) is essential for lymphocyte activation that includes cell proliferation and differentiation. This increase in [Ca2+]i results from Ca2+ release from the intracellular store and the subsequent Ca2+ influx from the extracellular environment via calcium channels located on the plasma membrane. Although transient receptor potential (TRP) channels have been reported to play important roles in the [Ca2+]i increase in lymphocytes, the function of these channels in lymphocyte activation remains unknown. Here, we report the comprehensive expression profile of TRP channel gene families including TRPC, TRPV, and TRPM in the murine immune system. RT-PCR analysis revealed different expression patterns of the TRP channel genes in B and T lymphocytes isolated from the spleen. Therefore, our results provide an appropriate reference of TRP gene expression in murine lymphocytes

  12. Soluble CD83 ameliorates experimental colitis in mice.

    Eckhardt, J; Kreiser, S; Döbbeler, M; Nicolette, C; DeBenedette, M A; Tcherepanova, I Y; Ostalecki, C; Pommer, A J; Becker, C; Günther, C; Zinser, E; Mak, T W; Steinkasserer, A; Lechmann, M

    2014-07-01

    The physiological balance between pro- and anti-inflammatory processes is dysregulated in inflammatory bowel diseases (IBD) as in Crohn's disease and ulcerative colitis. Conventional therapy uses anti-inflammatory and immunosuppressive corticosteroids to treat acute-phase symptoms. However, low remission rate and strong side effects of these therapies are not satisfying. Thus, there is a high medical need for new therapeutic strategies. Soluble CD83, the extracellular domain of the transmembrane CD83 molecule, has been reported to have interesting therapeutic and immunosuppressive properties by suppressing dendritic cell (DC)-mediated T-cell activation and inducing tolerogenic DCs. However, the expression and function of CD83 in IBD is still unknown. Here, we show that CD83 expression is upregulated by different leukocyte populations in a chemical-induced murine colitis model. Furthermore, in this study the potential of sCD83 to modulate colitis using an experimental murine colitis model was investigated. Strikingly, sCD83 ameliorated the clinical disease symptoms, drastically reduced mortality, and strongly decreased inflammatory cytokine expression in mesenteric lymph nodes and colon. The infiltration of macrophages and granulocytes into colonic tissues was vigorously inhibited. Mechanistically, we could show that sCD83-induced expression of indolamine 2,3-dioxygenase is essential for its protective effects. PMID:24424524

  13. Macrophage polarization in experimental and clinical choroidal neovascularization

    Yang, Yu; Liu, Fang; Tang, Miao; Yuan, Miner; Hu, Andina; Zhan, Zongyi; Li, Zijing; Li, Jiaqing; Ding, Xiaoyan; Lu, Lin

    2016-01-01

    Macrophages play an important role in the development of age-related macular degeneration (AMD). In this study, the spatial and temporal changes and the polarization of macrophages in murine laser-induced choroidal neovascularization (CNV) were investigated, and the polarized M1 and M2 biomarkers in the aqueous humors of neovascular AMD (nAMD) patients were studied. Macrophages, the main infiltrating inflammatory cells in CNV lesions, were evidenced by a significant increase in F4/80 mRNA expression and by the infiltration of F4/80+ cells in the lesions and the vicinity of laser-induced CNV. The mRNA expressions of M1-related markers were dramatically upregulated in the early stage, while the M2-related markers were slightly upregulated in the middle stage and sustained until the late stage. The results of immunostaining showed a similar early-but-transient M1 pattern and a delayed-but-sustained M2 pattern in laser-induced CNV. In addition, a higher M2/M1 ratio was found in both the murine models (Arg-1/iNOS and CCL22/CXCL10) and the aqueous humors of nAMD patients (CCL22/CXCL10) than in the controls. Our results suggested that the dynamic patterns of M1 and M2 were different in both the experimental and clinical CNV. The M2 macrophages were predominant and may play a more important role in the development of CNV. PMID:27489096

  14. Murine lymphoid procoagulant activity induced by bacterial lipopolysaccharide and immune complexes is a monocyte prothrombinase

    Schwartz, BS; Levy, GA; Fair, DS; Edgington, TS

    1982-01-01

    Murine lymphoid cells respond rapidly to bacterial lipopolysaccharide or antigen-antibody complexes to initiate or accelerate the blood coagulation pathways. The monocyte or macrophage has been identified as the cellular source, although lymphocyte collaboration is required for the rapid induction of the procoagulant response. This procoagulant activity is identified in the present study as a direct prothrombin activator, i.e., a prothrombinase. Studies with plasmas deficient in single coagul...

  15. Calcium regulates the commitment of murine erythroleukemia cells to terminal erythroid differentiation

    1981-01-01

    An alteration in the rate of calcium transport appears to be the rate- limiting event for the commitment of murine erythroleukemia (MEL) cells to initiate a program of terminal erythroid differentiation. The dimethyl sulfoxide (DMSO)-induced commitment of MEL cells to erythroid differentiation can be inhibited by treatment of cells with the calcium- chelating agent EGTA. Upon removal of EGTA, cells initiate commitment without the 12-h lag normally observed after treatment with DMSO alone. Tre...

  16. Prefoldin 5 Is Required for Normal Sensory and Neuronal Development in a Murine Model*

    Lee, Yongsuk; Smith, Richard S; Jordan, Wanda; Benjamin L. King; Won, Jungyeon; Valpuesta, Jose M.; Naggert, Jurgen K.; Nishina, Patsy M.

    2010-01-01

    Molecular chaperones and co-chaperones are crucial for cellular development and maintenance as they assist in protein folding and stabilization of unfolded or misfolded proteins. Prefoldin (PFDN), a ubiquitously expressed heterohexameric co-chaperone, is necessary for proper folding of nascent proteins, in particular, tubulin and actin. Here we show that a genetic disruption in the murine Pfdn5 gene, a subunit of prefoldin, causes a syndrome characterized by photoreceptor degeneration, centra...

  17. Aspirin inhibits in vitro maturation and in vivo immunostimulatory function of murine myeloid dendritic cells

    Hackstein, H; Morelli, AE; Larregina, AT; Ganster, RW; Papworth, GD; Logar, AJ; Watkins, SC; Falo, LD; Thomson, AW

    2001-01-01

    Aspirin is the most commonly used analgesic and antiinflammatory agent. In this study, at physiological concentrations, it profoundly inhibited CD40, CD80, CD86, and MHC class II expression on murine, GM-CSF + IL-4 stimulated, bone marrow-derived myeloid dendritic cells (DC). CD11c and MHC class I expression were unaffected. The inhibitory action was dose dependent and was evident at concentrations higher than those necessary to inhibit PG synthesis. Experiments with indomethacin revealed tha...

  18. Ovarian Aging-Like Phenotype in the Hyperandrogenism-Induced Murine Model of Polycystic Ovary

    Mohammad Amin Rezvanfar; Shojaei Saadi, Habib A; Maziar Gooshe; Amir Hosein Abdolghaffari; Maryam Baeeri; Mohammad Abdollahi

    2014-01-01

    There are prominently similar symptoms, effectors, and commonalities in the majority of characteristics between ovarian aging and polycystic ovarian syndrome (PCOS). Despite the approved role of oxidative stress in the pathogenesis of PCOS and aging, to our knowledge, the link between the PCO(S) and aging has not been investigated yet. In this study we investigated the possible exhibition of ovarian aging phenotype in murine model of PCO induced by daily oral administration of letrozole (1 mg...

  19. Adapting Human Videofluoroscopic Swallow Study Methods to Detect and Characterize Dysphagia in Murine Disease Models

    Lever, Teresa E.; Braun, Sabrina M.; Brooks, Ryan T.; Harris, Rebecca A.; Littrell, Loren L.; Neff, Ryan M.; Hinkel, Cameron J.; Allen, Mitchell J.; Ulsas, Mollie A.

    2015-01-01

    This study adapted human videofluoroscopic swallowing study (VFSS) methods for use with murine disease models for the purpose of facilitating translational dysphagia research. Successful outcomes are dependent upon three critical components: test chambers that permit self-feeding while standing unrestrained in a confined space, recipes that mask the aversive taste/odor of commercially-available oral contrast agents, and a step-by-step test protocol that permits quantification of swallow physi...

  20. Beneficial effect of Punica granatum peel extract on murine malaria-induced spleen injury

    Mubaraki, Murad A.; Hafiz, Taghreed A.; Dkhil, Mohamed A.; Al-Quraishy, Saleh

    2016-01-01

    Background Multiple drug-resistant malaria parasites have been widely detected, which has encouraged research studies focused on discovering alternative therapies. Medicinal plants such as pomegranate, Punica granatum, have been proven to exhibit antiprotozoal effects and therefore, we examined its effects on murine malaria-induced splenic injury and oxidative stress in this study. Methods Mice were divided into three groups, a vehicle control and two groups that were infected with 106 Plasmo...

  1. Dual Transgene Expression in Murine Cerebellar Purkinje Neurons by Viral Transduction In Vivo

    Bosch, Marie K.; Nerbonne, Jeanne M.; Ornitz, David M.

    2014-01-01

    Viral-vector mediated gene transfer to cerebellar Purkinje neurons in vivo is a promising avenue for gene therapy of cerebellar ataxias and for genetic manipulation in functional studies of animal models of cerebellar disease. Here, we report the results of experiments designed to identify efficient methods for viral transduction of adult murine Purkinje neurons in vivo. For these analyses, several lentiviral and an adeno-associated virus (AAV), serotype 1, vector with various promoter combin...

  2. Roles of interferon-gamma and interleukin-4 in murine lupus.

    Peng, S L; Moslehi, J; Craft, J.

    1997-01-01

    The systemic autoimmune syndrome of MRL/Mp-lpr/lpr (MRL/lpr) mice consists of severe pan-isotype hypergammaglobulinemia, autoantibody production, lymphadenopathy, and immune complex-associated end-organ disease. Its pathogenesis has been largely attributed to helper alphabeta T cells that may require critical cytokines to propagate pathogenic autoantibody production. To investigate the roles of prototypical Th1 and Th2 cytokines in the pathogenesis of murine lupus, IFN-gamma -/- and IL-4 -/- ...

  3. Protein stabilization explains the gag requirement for transformation of lymphoid cells by Abelson murine leukemia virus

    Prywes, R; Hoag, J; Rosenberg, N; Baltimore, D

    1985-01-01

    The single protein encoded by Abelson murine leukemia virus is a fusion of sequence from the retroviral gag genes with the v-abl sequence. Deletion of most of the gag region from the transforming protein results in a virus capable of transforming fibroblasts but no longer capable of transforming lymphoid cells. Smaller deletions in gag reveal that p15 gag sequences are responsible for this effect, whereas deletion of p12 sequences had no effect on lymphoid transformation. In transformed fibro...

  4. A novel mechanism of murine hepatocyte death inducible by Concanavalin A

    Leist, Marcel; Wendel, Albrecht

    1996-01-01

    Background: Concanavalin A (Con A) is a plant lectin that polyclonally activates T-cells. When given intravenously to mice it induces a selective liver failure. Hepatotoxicity following Con A administration involves the systemic release of tumor necrosis factor.Methods: We used primary murine hepatocyte cultures to investigate mechanisms of hepatocytotoxicity related to this animal model of inflammatory liver failure.Results: Con A was directly toxic for cultured hepatocytes. This toxicity di...

  5. Pharmacokinetic-Pharmacodynamic Analysis of Spiroindolone Analogs and KAE609 in a Murine Malaria Model

    Lakshminarayana, Suresh B.; Freymond, Céline; Fischli, Christoph; Yu, Jing; Weber, Sebastian; Goh, Anne; Yeung, Bryan K. S.; Ho, Paul C.; Dartois, Véronique; Diagana, Thierry T.; Rottmann, Matthias; Blasco, Francesca

    2014-01-01

    Limited information is available on the pharmacokinetic (PK) and pharmacodynamic (PD) parameters driving the efficacy of antimalarial drugs. Our objective in this study was to determine dose-response relationships of a panel of related spiroindolone analogs and identify the PK-PD index that correlates best with the efficacy of KAE609, a selected class representative. The dose-response efficacy studies were conducted in the Plasmodium berghei murine malaria model, and the relationship between ...

  6. Structural and biomechanical responses of osseous healing: a novel murine nonunion model

    Chaubey, Aditya; Grawe, Brian; Jeffrey A Meganck; Dyment, Nathaniel; Inzana, Jason; Jiang, Xi; Connolley, Camille; Awad, Hani; Rowe, David; Kenter, Keith; Goldstein, Steven A.; Butler, David

    2013-01-01

    Background Understanding the biological mechanisms of why certain fractures are at risk for delayed healing or nonunion requires translational animal models that take advantage of transgenic and other genetic manipulation technologies. Reliable murine nonunion models can be an important tool to understand the biology of nonunion. In this study, we report the results of a recently established model for creating critical defects that lead to atrophic nonunions based on a unique fracture fixatio...

  7. Development of a Murine Mycobacterial Growth Inhibition Assay for Evaluating Vaccines against Mycobacterium tuberculosis▿ †

    Parra, Marcela; Yang, Amy L.; Lim, Jaehyun; Kolibab, Kristopher; Derrick, Steven; Cadieux, Nathalie; Perera, Liyanage P.; Jacobs, William R.; Brennan, Michael; Morris, Sheldon L.

    2009-01-01

    The development and characterization of new tuberculosis (TB) vaccines has been impeded by the lack of reproducible and reliable in vitro assays for measuring vaccine activity. In this study, we developed a murine in vitro mycobacterial growth inhibition assay for evaluating TB vaccines that directly assesses the capacity of immune splenocytes to control the growth of Mycobacterium tuberculosis within infected macrophages. Using this in vitro assay, protective immune responses induced by immu...

  8. ABC- and SLC-Transporters in Murine and Bovine Mammary Epithelium - Effects of Prochloraz

    Yagdiran, Yagmur; Oskarsson, Agneta; Knight, Christopher H.; Tallkvist, Jonas

    2016-01-01

    Some chemicals are ligands to efflux transporters which may result in high concentrations in milk. Limited knowledge is available on the influence of maternal exposure to chemicals on the expression and function of transporters in the lactating mammary gland. We determined gene expression of ABC and SLC transporters in murine mammary tissue of different gestation and lactation stages, in murine mammary cells (HC11) featuring resting and secreting phenotypes and in bovine mammary tissue and cells (BME-UV). Effects on transporter expression and function of the imidazole fungicide prochloraz, previously reported to influence BCRP in mammary cells, was investigated on transporter expression and function in the two cell lines. Transporters studied were BCRP, MDR1, MRP1, OATP1A5/OATP1A2, OCTN1 and OCT1. Gene expressions of BCRP and OCT1 in murine mammary glands were increased during gestation and lactation, whereas MDR1, MRP1, OATP1A5 and OCTN1 were decreased, compared to expressions in virgins. All transporters measured in mammary glands of mice were detected in bovine mammary tissue and in HC11 cells, while only MDR1 and MRP1 were detected in BME-UV cells. Prochloraz treatment induced MDR1 gene and protein expression in both differentiated HC11 and BME-UV cells and increased protein function in HC11 cells, resulting in decreased accumulation of the MDR1 substrate digoxin. In conclusion, our results demonstrate that murine (HC11) and bovine (BME-UV) mammary epithelial cells can be applied to characterize expression and function of transporters as well as effects of contaminants on the mammary transporters. An altered expression, induced by a drug or toxic chemical, on any of the transporters expressed in the mammary epithelial cells during lactation may modulate the well-balanced composition of nutrients and/or secretion of contaminants in milk with potential adverse effects on breast-fed infants and dairy consumers. PMID:27028005

  9. Membrane-associated phosphoproteins in Plasmodium berghei-infected murine erythrocytes

    1983-01-01

    Normal and Plasmodium berghei (NYU-2 strain)-infected murine erythrocytes display substantially different patterns of plasma membrane phosphoproteins phosphorylation. Intact erythrocytes (normal and parasite infected) incubated with 32Pi and isolated washed erythrocyte plasma membranes incubated with gamma-32P-ATP were analyzed for phosphoproteins by SDS PAGE and autoradiography. Two new phosphoproteins of molecular weight 45,000 (pp45) and 68,000 (pp68), which are absent in normal erythrocyt...

  10. Expression of Adrenergic and Cholinergic Receptors in Murine Renal Intercalated Cells

    JUN, Jin-Gon; MAEDA, Seishi; Kuwahara-Otani, Sachi; TANAKA, Koichi; Hayakawa, Tetsu; SEKI, Makoto

    2014-01-01

    ABSTRACT Neurons influence renal function and help to regulate fluid homeostasis, blood pressure and ion excretion. Intercalated cells (ICCs) are distributed throughout the renal collecting ducts and help regulate acid/base equilibration. Because ICCs are located among principal cells, it has been difficult to determine the effects that efferent nerve fibers have on this cell population. In this study, we examined the expression of neurotransmitter receptors on the murine renal epithelial M-1...

  11. Effects of 810-nm Laser on Murine Bone-Marrow-Derived Dendritic Cells

    Chen, Aaron C.-H.; Huang, Ying-Ying; Sharma, Sulbha K.; Hamblin, Michael R.

    2011-01-01

    Objective: The purpose of this study was to Investigate the effect of 810-nm low level laser therapy (LLLT) on dendritic cells (DC) in vitro. Background data: LLLT can enhance wound healing and increase cell proliferation and survival, and is used to treat inflammatory conditions. However there are reports that LLLT can stimulate leukocytes and could therefore be pro-inflammatory. Recently, DC have been found to play an important role in inflammation and immune response. Methods: Murine bone-...

  12. Interaction of the Murine Dilute Suppressor Gene (Dsu) with Fourteen Coat Color Mutations

    Moore, K. J.; Swing, D. A.; Copeland, N. G.; Jenkins, N. A.

    1990-01-01

    The murine dilute suppressor gene, dsu, was previously shown to suppress the dilute coat color phenotypes of mice homozygous for the dilute (d), leaden (ln), and ashen (ash) mutations. Each of these mutations produce adendritic melanocytes, which results in an abnormal transportation of pigment granules into the hair shaft and a diluted coat color. The suppression of each mutation is associated with the restoration of near normal melanocyte morphology, indicating that dsu can compensate for t...

  13. Inflammatory and Remodeling Events in Asthma with Chronic Exposure to House Dust Mites: A Murine Model

    Ahn, Joong Hyun; Kim, Chi Hong; Kim, Yong Hyun; Kim, Seung Joon; Lee, Sook-Young; Kim, Young Kyoon; Kim, Kwan Hyoung; Moon, Hwa Sik; Song, Jeong Sup; Park, Sung Hak; Kwon, Soon Seog

    2007-01-01

    Although animal models with ovalbumin have been used to study chronic asthma, there are difficulties in inducing recurrence as well as in maintaining chronic inflammation in this system. Using a murine model of house dust mite (HDM)-induced bronchial asthma, we examined the airway remodeling process in response to the chronic exposure to HDM. During the seventh and twelfth weeks of study, HDM were inhaled through the nose for three consecutive days and airway responsiveness was measured. Twen...

  14. An increase in telomere sister chromatid exchange in murine embryonic stem cells possessing critically shortened telomeres

    Wang, Yisong; Erdmann, Natalie; Giannone, Richard J.; Wu, Jun; Gomez, Marla; Liu, Yie

    2005-01-01

    Telomerase deficiency leads to a progressive loss of telomeric DNA that eventually triggers cell apoptosis in human primary cells during prolonged growth in culture. Rare survivors can maintain telomere length through either activation of telomerase or recombination-based telomere lengthening, and thus proliferate indefinitely. We have explored the possibility that telomeres may be maintained through telomere sister chromatid exchange (T-SCE) in murine telomere reverse transcriptase-deficient...

  15. Cell surface polypeptides of murine T-cell clones expressing cytolytic or amplifier activity.

    Sarmiento, M.; Glasebrook, A L; Fitch, F. W.

    1980-01-01

    Murine cytolytic T-cell and amplifier T-cell clones derived from secondary unidirectional mixed leukocyte cultures were labeled with 125I by the lactoperoxidase method and their polypeptide profiles were analyzed by NaDodSO4/polyacrylamide gel electrophoresis. All cytolytic T-cell clones derived from the same mouse strain yeilded similar cell surface polypeptide profiles. However, profiles obtained with three amplifier T-cell clones were strikingly different from each other as well as from th...

  16. Methyltransferase G9A regulates T cell differentiation during murine intestinal inflammation

    Antignano, Frann; Burrows, Kyle; Hughes, Michael R.; Han, Jonathan M.; Kron, Ken J.; Penrod, Nadia M.; Oudhoff, Menno J.; Wang, Steven Kai Hao; Min, Paul H.; Gold, Matthew J; Chenery, Alistair L.; Braam, Mitchell J.S.; Fung, Thomas C.; Rossi, Fabio M.V.; McNagny, Kelly M.

    2014-01-01

    Inflammatory bowel disease (IBD) pathogenesis is associated with dysregulated CD4+ Th cell responses, with intestinal homeostasis depending on the balance between IL-17–producing Th17 and Foxp3+ Tregs. Differentiation of naive T cells into Th17 and Treg subsets is associated with specific gene expression profiles; however, the contribution of epigenetic mechanisms to controlling Th17 and Treg differentiation remains unclear. Using a murine T cell transfer model of colitis, we found that T cel...

  17. Cloning and Mutagenesis of the Murine Gammaherpesvirus 68 Genome as an Infectious Bacterial Artificial Chromosome

    Adler, Heiko; Messerle, Martin; Wagner, Markus; Koszinowski, Ulrich H.

    2000-01-01

    Gammaherpesviruses cause important infections of humans, in particular in immunocompromised patients. Recently, murine gammaherpesvirus 68 (MHV-68) infection of mice has been developed as a small animal model of gammaherpesvirus pathogenesis. Efficient generation of mutants of MHV-68 would significantly contribute to the understanding of viral gene functions in virus-host interaction, thereby further enhancing the potential of this model. To this end, we cloned the MHV-68 genome as a bacteria...

  18. The Effect of Erythropoietin on Neurotrophic Factors in N9 Murine Microglial Cells

    Kuralay, Filiz; ÇAKIRLI, Başak BİNGOL; GENÇ, Şermin

    2008-01-01

    Aim: In this study, we investigated whether interferon gamma (IFNg), lipopolysaccharides (LPS) and amyloid beta (AMYb), as toxic stimulator agents, and erythropoietin (EPO), as a neurotrophic agent, have an effect on the production of the following neurotrophic factors in the N9 murine microglia cell line: neurotrophin 3 (NT3), neurotrophin 4 (NT4), and brain-derived neurotrophic factor (BDNF). Materials and Methods: Microglial cells were incubated with 50 μg/ml AMYb, or 1 _...

  19. Dietary mistletoe lectin supplementation and reduced growth of a murine non-Hodgkin lymphoma

    Pryme, I.F.; Bardocz, S; Pusztai, A.; Ewen, S.W.B.

    2002-01-01

    The growth of a murine non-Hodgkin lymphoma (NHL) tumour has been shown to be reduced by incorporating mistletoe lectin (ML-1) into the diet. The morphological characteristics of NHL tumours in mice fed ML-1-supplemented diets were different from those in LA (control)-fed mice. The degree of mitotic activity was lower and nuclear area reduced. The degree of lymphocyte infiltration was increased in tumours from ML-1 fed mice and this was accompanied by a hig...

  20. Role of desensitization and subunit expression for kainate receptor-mediated neurotoxicity in murine neocortical cultures

    Jensen, J B; Schousboe, A; Pickering, D S;

    1999-01-01

    The neurotoxic actions of kainate and domoate were studied in cultured murine neocortical neurons at various days in culture and found to be developmentally regulated involving three components of neurotoxicity: (1) toxicity via indirect activation of N-methyl-D-aspartate (NMDA) receptors, (2) to...... produced by kainate receptors in mature cultures. Examining the subunit expression of the kainate receptor subunits GluR6/7 and KA2 did, however, not reveal any major change during development of the cultures....

  1. Bromelain Inhibits Allergic Sensitization and Murine Asthma via Modulation of Dendritic Cells

    Secor, Eric R.; Szczepanek, Steven M.; Castater, Christine A.; Adami, Alexander J.; Matson, Adam P.; Rafti, Ektor T.; Linda Guernsey; Prabitha Natarajan; McNamara, Jeffrey T.; Craig M. Schramm; Thrall, Roger S.; Silbart, Lawrence K.

    2013-01-01

    The incidence of atopic conditions has increased in industrialized countries. Persisting symptoms and concern for drug side-effects lead patients toward adjunctive treatments such as phytotherapy. Previously, we have shown that Bromelain (sBr), a mixture of cysteine proteases from pineapple, Ananas comosus, inhibits ovalbumin (OVA)-induced murine model of allergic airway disease (AAD). However, sBr's effect on development of AAD when treatment is administered throughout OVA-alum sensitization...

  2. Isolation and characterisation of Ebolavirus-specific recombinant antibody fragments from murine and shark immune libraries.

    Goodchild, Sarah A; Dooley, Helen; Schoepp, Randal J; Flajnik, Martin; Lonsdale, Stephen G

    2011-09-01

    Members of the genus Ebolavirus cause fulminating outbreaks of disease in human and non-human primate populations with a mortality rate up to 90%. To facilitate rapid detection of these pathogens in clinical and environmental samples, robust reagents capable of providing sensitive and specific detection are required. In this work recombinant antibody libraries were generated from murine (single chain variable domain fragment; scFv) and nurse shark, Ginglymostoma cirratum (IgNAR V) hosts immunised with Zaire ebolavirus. This provides the first recorded IgNAR V response against a particulate antigen in the nurse shark. Both murine scFv and shark IgNAR V libraries were panned by phage display technology to identify useful antibodies for the generation of immunological detection reagents. Two murine scFv were shown to have specificity to the Zaire ebolavirus viral matrix protein VP40. Two isolated IgNAR V were shown to bind to the viral nucleoprotein (NP) and to capture viable Zaire ebolavirus with a high degree of sensitivity. Assays developed with IgNAR V cross-reacted to Reston ebolavirus, Sudan ebolavirus and Bundibugyo ebolavirus. Despite this broad reactivity, neither of IgNAR V showed reactivity to Côte d'Ivoire ebolavirus. IgNAR V was substantially more resistant to irreversible thermal denaturation than murine scFv and monoclonal IgG in a comparative test. The demonstrable robustness of the IgNAR V domains may offer enhanced utility as immunological detection reagents in fieldable biosensor applications for use in tropical or subtropical countries where outbreaks of Ebolavirus haemorrhagic fever occur. PMID:21752470

  3. Antiviral Effects of Antisense Morpholino Oligomers in Murine Coronavirus Infection Models▿

    Burrer, Renaud; Neuman, Benjamin W.; Ting, Joey P.C.; Stein, David A.; Moulton, Hong M.; Iversen, Patrick L.; Kuhn, Peter; Michael J Buchmeier

    2007-01-01

    The recent emergence of novel pathogenic human and animal coronaviruses has highlighted the need for antiviral therapies that are effective against a spectrum of these viruses. We have used several strains of murine hepatitis virus (MHV) in cell culture and in vivo in mouse models to investigate the antiviral characteristics of peptide-conjugated antisense phosphorodiamidate morpholino oligomers (P-PMOs). Ten P-PMOs directed against various target sites in the viral genome were tested in cell...

  4. Connexin43 Cardiac Gap Junction Remodeling: Lessons from Genetically Engineered Murine Models

    Remo, Benjamin F.; Giovannone, Steven; Fishman, Glenn I.

    2012-01-01

    Sudden cardiac death is responsible for several hundred thousand deaths each year in the United States. Multiple lines of evidence suggest that perturbation of gap junction expression and function in the heart, or what has come to be known as cardiac gap junction remodeling, plays a key mechanistic role in the pathophysiology of clinically significant cardiac arrhythmias. Here we review recent studies from our laboratory using genetically engineered murine models to explore mechanisms implica...

  5. Effect of Tityus serrulatus venom on cytokine production and the activity of murine macrophages

    Vera L. Petricevich

    2002-01-01

    THE purpose of this study was to investigate the effects of Tityus serrulatus venom (TSV) on murine peritoneal macrophages evaluated in terms of activation. The effects of crude TSV were analysed by detection of cytokines, oxygen intermediate metabolites (H2O2) and nitric oxide (NO) in supernatants of peritoneal macrophages. Several functional bioassays were employed including an in vitro model for envenomating: cytotoxicity of TSV was assessed using the lyses percentage. Tumor necrosis facto...

  6. Interactome analysis of myeloid-derived suppressor cells in murine models of colon and breast cancer

    Aliper, Alexander M.; FRIEDEN-KOROVKINA, VICTORIA P.; Buzdin, Anton; Roumiantsev, Sergey A.; Zhavoronkov, Alex

    2014-01-01

    In solid cancers, myeloid derived suppressor cells (MDSC) infiltrate (peri)tumoral tissues to induce immune tolerance and hence to establish a microenvironment permissive to tumor growth. Importantly, the mechanisms that facilitate such infiltration or a subsequent immune suppression are not fully understood. Hence, in this study, we aimed to delineate disparate molecular pathways which MDSC utilize in murine models of colon or breast cancer. Using pathways enrichment analysis, we completed i...

  7. Review: 2-mercaptoethanol alteration of in vitro immune functions of species other than murine

    Click, Robert E

    2013-01-01

    Descriptions that organosulfurs could alter biologically relevant cellular functions began some 40 years ago when cell mediated and humoral murine in vitro immune responses were reported to be dramatically enhanced by any of four xenobiotic, sulfhydryl compounds—2-mercaptoethanol (2-ME), dithiothreitol, glutathione, and L-cysteine; the most effective of the four was 2-ME. These findings triggered a plethora of reports defining 2-ME benefits for a multitude of immunological processes, primaril...

  8. Increased Resistance to Taenia crassiceps Murine Cysticercosis in Qa-2 Transgenic Mice

    Fragoso, Gladis; Lamoyi, Edmundo; Mellor, Andrew; Lomelí, Ciro; Hernández, Marisela; Sciutto, Edda

    1998-01-01

    We previously reported important differences in resistance to Taenia crassiceps murine cysticercosis between BALB/c substrains. It was suggested that resistance might correlate with expression of the nonclassic class I major histocompatibility complex (MHC) Qa-2 antigen; BALB/cAnN is Qa-2 negative and highly susceptible to T. crassiceps, whereas BALB/cJ expresses Qa-2 and is highly resistant. In this study, we investigated the role of Qa-2 in mediating resistance to cysticercosis by linkage a...

  9. In Vivo Pharmacokinetics and Pharmacodynamics of a New Triazole, Voriconazole, in a Murine Candidiasis Model

    Andes, D.; Marchillo, K.; Stamstad, T.; Conklin, R.

    2003-01-01

    In vivo studies have described the pharmacodynamic (PD) characteristics of several triazoles. These investigations have demonstrated that the 24-h area under the concentration-time curve (AUC)/MIC ratio is the critical pharmacokinetic (PK)-PD parameter associated with treatment efficacy. Further analyses from these in vivo studies have demonstrated that a triazole free drug 24-h AUC/MIC of 20 to 25 is predictive of treatment success. We used a neutropenic murine model of disseminated Candida ...

  10. Absence of proteins related to murine mammary tumour virus polypeptides in rat mammary tumours

    Since normal rat DNA contains sequences which hybridize with the genome of the murine mammary tumour virus (MuMTV), it is possible that a related virus would play a role in mammary carcinogenesis in rats. The authors screened a number of rat mammary tumours for antigens related to the MuMTV polypeptides gp52 and p28 by means of a radioimmunoassay. (Auth.)

  11. Inhibition of inducible Nitric Oxide Synthase by a mustard gas analog in murine macrophages

    Smith Milton; Yang Hongsong; Paromov Victor M; Qui Min; Stone William L

    2006-01-01

    Abstract Background 2-Chloroethyl ethyl sulphide (CEES) is a sulphur vesicating agent and an analogue of the chemical warfare agent 2,2'-dichlorodiethyl sulphide, or sulphur mustard gas (HD). Both CEES and HD are alkylating agents that influence cellular thiols and are highly toxic. In a previous publication, we reported that lipopolysaccharide (LPS) enhances the cytotoxicity of CEES in murine RAW264.7 macrophages. In the present investigation, we studied the influence of CEES on nitric oxide...

  12. Aging Leads to a Programmed Loss of Brown Adipocytes in Murine Subcutaneous White Adipose Tissue

    Rogers, Nicole H; Landa, Alejandro; Park, Seongjoon; Smith, Roy G.

    2012-01-01

    Insulin sensitivity deteriorates with age, but mechanisms remain unclear. Age-related changes in the function of subcutaneous white adipose tissue (sWAT) are less characterized than those in visceral WAT. We hypothesized that metabolic alterations in sWAT, which in contrast to epididymal WAT, harbors a sub-population of energy dissipating UCP1+ brown adipocytes, promote age-dependent progression towards insulin resistance. Indeed, we show that a predominant consequence of aging in murine sWAT...

  13. Regulation of murine macrophage Ia-antigen expression by products of activated spleen cells

    1980-01-01

    This investigation examined the effects of mediators derived form activated spleen cells on macrophage Ia-antigen expression and function. Incubation of adherent thioglycollate-induced murine peritoneal macrophages(> 90% Ia-) with concanavalin A (Con A)- stimulated spleen cell supernate (Con A sup) resulted in a dose- dependent increase in the percentage of Ia-containing (Ia+) phagocytic cells, as detected by antiserum-and-complement-mediated cytotoxicity. The Ia-antigen expression of macroph...

  14. Role of immunity in age-related resistance to paralysis after murine leukemia virus infection.

    Hoffman, P M; Robbins, D S; Morse, H C

    1984-01-01

    Resistance to the paralytic effects of a wild mouse (Cas-Br-M) murine leukemia virus infection develops with age and is complete by 10 days of age in susceptible NFS mice. The possibility that cell-mediated immunity plays a significant role in this resistance was suggested by the observation that treatment of 10-day-old mice with antithymocyte serum rendered them susceptible to paralysis. By comparison, mice rendered incapable of generating a humoral immune response by treatment from birth to...

  15. Dextrin nanoparticles : studies on the interaction with murine macrophages and blood clearance

    Gonçalves, Catarina; Torrado, Egídio; Martins, Teresa G.; Pereira, Paula; Pedrosa, Jorge; Gama, F. M.

    2010-01-01

    The uptake of nanoparticles by cells of the mononuclear phagocytic system limits its use as colloidal drug carriers, reducing the blood circulation time and the ability to reach biological targets. In this work, the interaction between dextrin nanoparticles – recently developed in our laboratory – and murine bone marrow-derived macrophages was evaluated. Cytotoxicity and nitric oxide production were studied, using the MTT assay and the Griess method, respectively. FITC labelled na...

  16. The Effect of Diphtheria Toxin on Nitric Oxide Induction From RAW264.7 Murine Macrophages

    Aybay, Cemalettin

    1998-01-01

    In this study the effect of diphtheria toxin (DT) on nitric oxide (NO) production from RAW 264.7 murine macrophages was investigated. Griess reagent was used to determine NO production by measuring nitrite levels in the culture supernatants. Corynebacterium diphtheriae G12/6 strain-produced DT (Limes flocculation activity and immunodiffusion assays were positive) demonstrated a dose-dependent effect on RAW 264.7 macrophages to induce NO production. L-NAME, an L-arginine analogue, ...

  17. A statistical analysis of murin incisional and excisional acute wound models.

    Ansell, David M; Campbell, Laura; Thomason, Helen A; Brass, Andrew; Hardman, Matthew J.

    2014-01-01

    Mice represent the most commonly used species for preclinical in vivo research. While incisional and excisional acute murine wound models are both frequently employed, there is little agreement on which model is optimum. Moreover, current lack of standardization of wounding procedure, analysis time point(s), method of assessment, and the use of individual wounds vs. individual animals as replicates makes it difficult to compare across studies. Here we have profiled secondary intention healing...

  18. A statistical analysis of murine incisional and excisional acute wound models

    Ansell, David M; Campbell, Laura; Thomason, Helen A; Brass, Andrew; Hardman, Matthew J.

    2014-01-01

    Mice represent the most commonly used species for preclinical in vivo research. While incisional and excisional acute murine wound models are both frequently employed, there is little agreement on which model is optimum. Moreover, current lack of standardization of wounding procedure, analysis time point(s), method of assessment, and the use of individual wounds vs. individual animals as replicates makes it difficult to compare across studies. Here we have profiled secondary intention healing...

  19. Killed Candida albicans Yeasts and Hyphae Inhibit Gamma Interferon Release by Murine Natural Killer Cells

    Murciano, Celia; Villamón, Eva; O'Connor, José-Enrique; Gozalbo, Daniel; Gil, M. Luisa

    2006-01-01

    Killed yeasts and hyphae of Candida albicans inhibit gamma interferon secretion by highly purified murine NK cells in response to the Toll-like receptor ligands lipopolysaccharide and zymosan. This effect, which is also observed in the presence of NK-activating cytokines (interleukin-2 [IL-2], IL-12, and IL-15), may represent a novel mechanism of immune evasion that contributes to the virulence of C. albicans.

  20. Killed Candida albicans yeasts and hyphae inhibit gamma interferon release by murine natural killer cells.

    Murciano, Celia; Villamón, Eva; O'Connor, José-Enrique; Gozalbo, Daniel; Gil, M Luisa

    2006-02-01

    Killed yeasts and hyphae of Candida albicans inhibit gamma interferon secretion by highly purified murine NK cells in response to the Toll-like receptor ligands lipopolysaccharide and zymosan. This effect, which is also observed in the presence of NK-activating cytokines (interleukin-2 [IL-2], IL-12, and IL-15), may represent a novel mechanism of immune evasion that contributes to the virulence of C. albicans. PMID:16428793

  1. The effect of folate on the methotrexate/indomethacin interaction in a murine cancer cell line.

    Hollingsworth, S J; Anderson, E. M.; Bennett, A

    1995-01-01

    1. The effect of folate on the interaction between methotrexate (a folate analogue) and indomethacin has been examined in murine NC carcinoma cells. 2. Conditioning of NC cells to a physiological (20 nM) folate concentration after culture in a high folate concentration increased the response to methotrexate. The sensitivity of these conditioned cells to methotrexate related inversely to the folate concentration. 3. At 20 nM and 2 microM folate, indomethacin 1 micrograms ml-1 potentiated the c...

  2. Protection against murine disseminated candidiasis mediated by a Candida albicans-specific T-cell line.

    Sieck, T G; Moors, M A; Buckley, H R; Blank, K J

    1993-01-01

    The role of T lymphocytes in disseminated candidiasis in a mouse model of irradiation-induced immunosuppression was investigated. A continuously cultured Candida albicans-specific T-cell line mediated protection of sublethally irradiated mice from disseminated candidiasis as measured by both the fungal load in the kidneys and mortality. These results are the first to demonstrate directly a role for antigen-specific T cells in the protective immune response against murine disseminated candidia...

  3. Production and Characterization of a Murine Monoclonal Antibody Against Human Ferritin

    Bayat, Ali Ahmad; Yeganeh, Omid; Ghods, Roya; Zarnani, Amir Hassan; Ardekani, Reza Bahjati; Mahmoudi, Ahmad Reza; Mahmoudian, Jafar; Haghighat-Noutash, Farzaneh; Jeddi-Tehrani, Mahmood

    2013-01-01

    Background Ferritin is an iron storage protein, which plays a key role in iron metabolism. Measurement of ferritin level in serum is one of the most useful indicators of iron status and also a sensitive measurement of iron deficiency. Monoclonal antibodies may be useful as a tool in various aspects of ferritin investigations. In this paper, the production of a murine monoclonal antibody (mAb) against human ferritin was reported. Methods Balb/c mice were immunized with purified human ferritin ...

  4. Functional dissection of the Moloney murine leukemia virus envelope protein gp70.

    Bae, Y.; Kingsman, S M; Kingsman, A J

    1997-01-01

    The envelope protein of Moloney murine leukemia virus (Mo-MLV) is a complex glycoprotein that mediates receptor binding and entry via fusion with cell membranes. By using a series of substitution mutations and truncations in the Mo-MLV external envelope surface protein gp70, we have identified regions important for these processes. Firstly, truncations of gp70 revealed that the minimal continuous receptor-binding region is amino acids 9 to 230, in broad agreement with other studies. Secondly,...

  5. Cloning and expression of murine enzymes involved in the salvage pathway of GDP-L-fucose.

    Niittymäki, Jaana; Mattila, Pirkko; Roos, Christophe; Huopaniemi, Laura; Sjöblom, Solveig; Renkonen, Risto

    2004-01-01

    In the salvage pathway of GDP-L-fucose, free cytosolic fucose is phosphorylated by L-fucokinase to form L-fucose-L-phosphate, which is then further converted to GDP-L-fucose in the reaction catalyzed by GDP-L-fucose pyrophosphorylase. We report here the cloning and expression of murine L-fucokinase and GDP-L-fucose pyrophosphorylase. Murine L-fucokinase is expressed as two transcripts of 3057 and 3270 base pairs, encoding proteins of 1019 and 1090 amino acids with predicted molecular masses of 111 kDa and 120 kDa respectively. Only the longer splice variant of L-fucokinase was enzymatically active when expressed in COS-7 cells. Murine GDP-L-fucose pyrophosphorylase has an open reading frame of 1773 base pairs encoding a protein of 591 amino acids with a predicted molecular mass of 65.5 kDa. GDP-L-fucose, the reaction product of GDP-L-pyrophosphorylase, was identified by HPLC and MALDI-TOF MS analysis. The tissue distribution of murine L-fucokinase and GDP-L-fucose pyrophosphorylase was investigated by quantitative real time PCR, which revealed high expression of L-fucokinase and GDP-L-fucose pyrophosphorylase in various tissues. The wide expression of both enzymes can also be observed from the large amount of data collected from a number of expressed sequence tag libraries, which indicate that not only the de novo pathway alone, but also the salvage pathway, could have a significant role in the synthesis of GDP-L-fucose in the cytosol. PMID:14686921

  6. Knockdown of the Placental Growth Factor Gene Inhibits Laser Induced Choroidal Neovascularization in a Murine Model

    Ramin Nourinia; Zahra-Soheila Soheili; Hamid Ahmadieh; Hassan Akrami; Mozhgan Rezaei Kanavi; Shahram Samiei

    2013-01-01

    Purpose: To evaluate the effect of placental growth factor (PlGF) gene knockdown in a murine model of laser-induced choroidal neovascularization. Methods: Choroidal neovascularization was induced in the left eyes of 11 mice by infrared laser. Small interfering RNA (siRNA, 20 picomoles/10 μl) corresponding to PlGF mRNA was administered intravitreally by Hamilton syringe in all subjects. One month later, fluorescein angiography and histolologic examination were performed. Results: No le...

  7. Alternans in genetically modified Langendorff-perfused murine hearts modeling catecholaminergic polymorphic ventricular tachycardia

    Ian N Sabir

    2010-10-01

    Full Text Available The relationship between alternans and arrhythmogenicity was studied in genetically modified murine hearts modeling catecholaminergic polymorphic ventricular tachycardia (CPVT during Langendorff perfusion, before and after treatment with catecholamines and a β-adrenergic antagonist. Heterozygous (RyR2p/s and homozygous (RyR2s/s RyR2-P2328S hearts, and wild-type (WT controls, were studied before and after treatment with epinephrine (100 nM and 1 µM and propranolol (100 nM. Monophasic action potential recordings demonstrated significantly greater incidences of arrhythmia in RyR2s/p and RyR2s/s hearts as compared to WTs. Arrhythmogenicity in RyR2s/s hearts was associated with alternans, particularly at short baseline cycle lengths. Both phenomena were significantly accentuated by treatment with epinephrine and significantly diminished by treatment with propranolol, in full agreement with clinical expectations. These changes took place, however, despite an absence of changes in action potential durations, ventricular effective refractory periods or restitution curve characteristics. Furthermore pooled data from all hearts in which arrhythmia occurred demonstrated significantly greater alternans magnitudes, but similar restitution curve slopes, to hearts that did not demonstrate arrhythmia. These findings thus further validate the RyR2-P2328S murine heart as a model for human CPVT, confirming an alternans phenotype in common with murine genetic models of the Brugada syndrome and the congenital long-QT syndrome type 3. In contrast to these latter similarities, however, this report demonstrates the dissociation of alternans from changes in the properties of restitution curves for the first time in a murine model of a human arrhythmic syndrome.

  8. Structure of the Murine Constitutive Androstane Receptor Complexed to Androstenol: A Molecular Basis for Inverse Agonism

    Shan, Li; Vincent, Jeremy; Brunzelle, Joseph S.; Dussault, Isabelle; Lin, Min; Ianculescu, Irina; Sherman, Mark A.; Forman, Barry M.; Fernandez, Elias J.

    2004-01-01

    The nuclear receptor CAR is a xenobiotic responsive transcription factor that plays a central role in the clearance of drugs and bilirubin while promoting cocaine and acetaminophen toxicity. In addition, CAR has established a “reverse” paradigm of nuclear receptor action where the receptor is active in the absence of ligand and inactive when bound to inverse agonists. We now report the crystal structure of murine CAR bound to the inverse agonist androstenol. Androstenol binds within the ligan...

  9. Murine Gammaherpesvirus 68 Evades Host Cytokine Production via Replication Transactivator-Induced RelA Degradation

    Dong, Xiaonan; He, Zhiheng; Durakoglugil, Deniz; Arneson, Lisa; Shen, Yan; Feng, Pinghui

    2012-01-01

    Cytokines play crucial roles in curtailing the propagation and spread of pathogens within the host. As obligate pathogens, gammaherpesviruses have evolved a plethora of mechanisms to evade host immune responses. We have previously shown that murine gammaherpesvirus 68 (γHV68) induces the degradation of RelA, an essential subunit of the transcriptionally active NF-κB dimer, to evade cytokine production. Here, we report that the immediately early gene product of γHV68, replication transactivato...

  10. Gene Therapy Prolongs Survival and Restores Function in Murine and Canine Models of Myotubular Myopathy

    Childers, Martin K.; Joubert, Romain; Poulard, Karine; Moal, Christelle; Grange, Robert W.; Doering, Jonathan A; Lawlor, Michael W.; Rider, Branden E; Jamet, Thibaud; Danièle, Nathalie; Martin, Samia; Rivière, Christel; Soker, Thomas; Hammer, Caroline; Van Wittenberghe, Laetitia

    2014-01-01

    Loss-of-function mutations in the myotubularin gene (MTM1) cause X-linked myotubular myopathy (XLMTM), a fatal, congenital pediatric disease that affects the entire skeletal musculature. Systemic administration of a single dose of a recombinant serotype-8 adeno-associated virus (AAV8) vector expressing murine myotubularin to Mtm1-deficient knockout mice at the onset or at late stages of the disease resulted in robust improvement in motor activity and contractile force, corrected muscle pathol...

  11. Ag85B DNA vaccine suppresses airway inflammation in a murine model of asthma

    Gao Xinglin

    2009-06-01

    Full Text Available Abstract Background In allergic asthma, Th2 lymphocytes are believed to play important roles in orchestrating airway eosinophilia and inflammation. Resetting the Th1/Th2 imbalance may have a therapeutic role in asthma. The mycobacterium tuberculosis 30-kilodalton major secretory protein (antigen 85B, Ag85B can protect animals from M. tuberculosis infection by inducing a Th1-dominant response. Methods In this study, the Ag85B gene was cloned into pMG plasmids to yield the pMG-Ag85B plasmid. The expression of Ag85B gene in murine bronchial epithelia cells was detected by Western blotting and immunohistochemical staining after intranasal immunization with reconstructed pMG-Ag85B plasmids. The protective effect of pMG-Ag85B plasmids immunization in airway inflammation was evaluated by histological examination and bronchoalveolar lavage (BAL. IL-4 and IFN-γ levels in the BAL and supernatant from splenocyte culture were determined using ELISA kits. Results The Ag85B gene was successfully expressed in murine bronchial epithelia cells by intranasal immunization with reconstructed pMG-Ag85B plasmids. Using a murine model of asthma induced by ovalbumin (OVA, pMG-Ag85B immunization significantly inhibited cellular infiltration across the airway epithelium with a 37% decrease in the total number of cells (9.6 ± 2.6 × 105/ml vs. 15.2 ± 3.0 × 105/ml, p 5/ml vs. 5.4 ± 1.1 × 105/ml, p Conclusion In a murine model of asthma induced by OVA, intranasal immunization with pMG-Ag85B significantly reduced allergic airway inflammation with less eosinophil infiltration. This protective effect was associated with decreased IL-4 and increased IFN-γ production in the BAL fluid and in the supernatant of cultured splenocytes.

  12. Antiproliferative effect of the jararhagin toxin on B16F10 murine melanoma

    Maria, Durvanei Augusto; da Silva, Manuela Garcia Laveli; Correia, Mario Cesar; Ruiz, Itamar Romano Garcia

    2014-01-01

    Background Malignant melanoma is a less common but highly dangerous form of skin cancer; it starts in the melanocytes cells found in the outer layer of the skin. Jararhagin toxin, a metalloproteinase isolated from Bothrops jararaca snake venom acts upon several biological processes, as inflammation, pain, platelet aggregation, proliferation and apoptosis, though not yet approved for use, may one day be employed to treat tumors. Methods B16F10 murine melanoma cells were treated with jararhagin...

  13. Homocysteine enhances MMP-9 production in murine macrophages via ERK and Akt signaling pathways

    Lee, Seung Jin; Lee, Yi Sle; Seo, Kyo Won; Bae, Jin Ung; Kim, Gyu Hee; Park, So Youn; Kim, Chi Dae, E-mail: chidkim@pusan.ac.kr

    2012-04-01

    Homocysteine (Hcy) at elevated levels is an independent risk factor of cardiovascular diseases, including atherosclerosis. In the present study, we investigated the effect of Hcy on the production of matrix metalloproteinases (MMP) in murine macrophages. Among the MMP known to regulate the activities of collagenase and gelatinase, Hcy exclusively increased the gelatinolytic activity of MMP-9 in J774A.1 cells as well as in mouse peritoneal macrophages. Furthermore, this activity was found to be correlated with Western blot findings in J774A.1 cells, which showed that MMP-9 expression was concentration- and time-dependently increased by Hcy. Inhibition of the ERK and Akt pathways led to a significant decrease in Hcy-induced MMP-9 expression, and combined treatment with inhibitors of the ERK and Akt pathways showed an additive effects. Activity assays for ERK and Akt showed that Hcy increased the phosphorylation of both, but these phosphorylation were not affected by inhibitors of the Akt and ERK pathways. In line with these findings, the molecular inhibition of ERK and Akt using siRNA did not affect the Hcy-induced phosphorylation of Akt and ERK, respectively. Taken together, these findings suggest that Hcy enhances MMP-9 production in murine macrophages by separately activating the ERK and Akt signaling pathways. -- Highlights: ► Homocysteine (Hcy) induced MMP-9 production in murine macrophages. ► Hcy induced MMP-9 production through ERK and Akt signaling pathways. ► ERK and Akt signaling pathways were activated by Hcy in murine macrophages. ► ERK and Akt pathways were additively act on Hcy-induced MMP-9 production. ► Hcy enhances MMP-9 production in macrophages via activation of ERK and Akt signaling pathways in an independent manner.

  14. Homocysteine enhances MMP-9 production in murine macrophages via ERK and Akt signaling pathways

    Homocysteine (Hcy) at elevated levels is an independent risk factor of cardiovascular diseases, including atherosclerosis. In the present study, we investigated the effect of Hcy on the production of matrix metalloproteinases (MMP) in murine macrophages. Among the MMP known to regulate the activities of collagenase and gelatinase, Hcy exclusively increased the gelatinolytic activity of MMP-9 in J774A.1 cells as well as in mouse peritoneal macrophages. Furthermore, this activity was found to be correlated with Western blot findings in J774A.1 cells, which showed that MMP-9 expression was concentration- and time-dependently increased by Hcy. Inhibition of the ERK and Akt pathways led to a significant decrease in Hcy-induced MMP-9 expression, and combined treatment with inhibitors of the ERK and Akt pathways showed an additive effects. Activity assays for ERK and Akt showed that Hcy increased the phosphorylation of both, but these phosphorylation were not affected by inhibitors of the Akt and ERK pathways. In line with these findings, the molecular inhibition of ERK and Akt using siRNA did not affect the Hcy-induced phosphorylation of Akt and ERK, respectively. Taken together, these findings suggest that Hcy enhances MMP-9 production in murine macrophages by separately activating the ERK and Akt signaling pathways. -- Highlights: ► Homocysteine (Hcy) induced MMP-9 production in murine macrophages. ► Hcy induced MMP-9 production through ERK and Akt signaling pathways. ► ERK and Akt signaling pathways were activated by Hcy in murine macrophages. ► ERK and Akt pathways were additively act on Hcy-induced MMP-9 production. ► Hcy enhances MMP-9 production in macrophages via activation of ERK and Akt signaling pathways in an independent manner.

  15. The genomic sequence of the murine major vault protein and its promoter.

    Mossink, Marieke; van Zon, Arend; Fränzel-Luiten, Erna; Schoester, Martijn; Scheffer, George L; Scheper, Rik J; Sonneveld, Pieter; Wiemer, Erik A C

    2002-07-10

    Vaults are ribonucleoproteins of unknown function, consisting of three different proteins and multiple copies of small untranslated RNA molecules. One of the protein subunits has been identified as TEP1, a protein that is also associated with the telomerase complex. Another protein appears to contain a functional PARP domain and is hence called VPARP. The third protein, major vault protein (MVP), is believed to make up 70% of the total mass of the vault complex and to be responsible for the typical barrel-shaped structure of vaults. We have isolated the murine MVP cDNA and compared the amino acid sequence with MVP from other species. Over 90% of sequence identity was found between mouse, human and rat, and a considerable degree of identity between mouse and MVPs from lower eukaryotes. We also found that the genomic structure of the murine MVP gene closely resembles the organization of the human MVP gene, both consisting of 15 exons of which most have exactly the same size. Finally we have isolated a genomic region upstream (and partially overlapping) the first untranslated exon, that displayed promoter activity in a luciferase reporter assay. Furthermore, we showed that the sequences from the first exon together with the 5'-end of the first intron enhance the promoter activity, implying the presence of essential promoter elements in this region. Alignment of the murine promoter region with the homologous sequences of the human gene revealed an identity of 58%. The apparent presence of conserved promoter elements suggests a similar regulation of human and murine MVP expression. PMID:12234684

  16. Alveolar macrophages modulate allergic inflammation in a murine model of asthma

    Bang, Bo-Ram; Chun, Eunyoung; Shim, Eun-Jin; Lee, Hyun-Seung; Lee, Soo-Yeon; Cho, Sang-Heon; Min, Kyung-Up; Kim, You-Young; Park, Heung-Woo

    2011-01-01

    The role of alveolar macrophages (AMs) in the pathogenesis of asthma is still unknown. The aim of the present study was to investigate the effects of AM in the murine model of asthma. AMs were selectively depleted by liposomes containing clodronate just before allergen challenges, and changes in inflammatory cells and cytokine concentrations in bronchoalveolar lavage (BAL) fluid were measured. AMs were then adoptively transferred to AM-depleted sensitized mice and changes were measured. Pheno...

  17. Correction of murine mucopolysaccharidosis VII by a human. beta. -glucuronidase transgene

    Kyle, J.W.; Vogler, C.; Hoffmann, J.W.; Sly, W.S. (St. Louis Univ. School of Medicine, MO (USA)); Birkenmeier, E.H.; Gwynn, B. (Jackson Laboratory, Bar Harbor, ME (USA))

    1990-05-01

    The authors recently described a murine model for mucopolysaccharidosis VII in mice that have an inherited deficiency of {beta}-glucuronidase. Affected mice, of genotype gus{sup mps}/gus{sup mps}, present clinical manifestations similar to those of humans with mucopolysaccharidosis VII (Sly syndrome) and are shown here to have secondary elevations of other lysosomal enzymes. The mucopolysaccharidosis VII phenotype in both species includes dwarfism, skeletal deformities, and premature death. Lysosome storage is visualized within enlarged vesicles and correlates biochemically with accumulation of undegraded and partially degraded glycosaminoglycans. In this report they describe the consequences of introducing the human {beta}-glucuronidase gene, GUSB, into gus{sup mps}/gus{sup mps} mice that produce virtually no murine {beta}-glucuronidase. Transgenic mice homozygous for the mucopolysaccharidosis VII mutation expressed high levels of human {beta}-glucuronidase activity in all tissues examined and were phenotypically normal. Biochemically, both the intralysosomal storage of glycosaminoglycans and the secondary elevation of other acid hydrolases were corrected. These findings demonstrate that the GUSB transgene is expressed in gus{sup mps}/gus{sup mps} mice and that human {beta}-glucuronidase corrects the murine mucopolysaccharidosis storage disease.

  18. Effect of small dose of radiation on induction of apoptosis in murine tumors

    To investigate the presence of adaptive response by low dose radiation in murine tumors in relation to radiation induced apoptosis as well as related mechanism. Syngeneic murine tumors, OCa-1 and HCa-l, were given 0.05 Gy pretreatment followed by therapeutic dose of 25 Gy radiation. Induction of apoptosis was analyzed for each treatment group. Regulating molecules of apoptosis. p53, Bcl-2, Sax, Bel-X, were also analyzed by Western blotting. In 0.05 Gy pretreatment group of OCa-l, 25 Gy-induced apoptosis per 1000 cells was 229, which was estimated at 30% lower level than the expected (p<0.05). In contrast, this reduction in radiation induced apoptosis was not seen in HCa-1. In the expression of apoptosis regulating molecules, p53 increased in both tumors in response to radiation. Bcl-2 and Bax did not show significant change in both tumors however, the expression of Bcl-2 surpassed that of Bax in 0.05 Gy pretreatment group of OCa-1. Bcl-X was not expressed in OCa-1. In HCa-l, ScI-X showed increased expression even with 0.05 Gy. Adaptive response by low dose radiation is shown in one murine tumor, OCa-I, in relation to radiation induced apoptosis. Apoptosis regulating molecules including Bcl-2/Bax and Bcl-X, appear to related. This study shows an evidence that adaptive response is present, but not a generalized phenomenon in vivo

  19. Alteration in transcription factor binding in murine thymocytes after low dose radiation

    Objective: To study the effect of low dose radiation on gene transcription regulation of murine thymocytes. Methods: Alteration in transcription factor binding in murine thymocytes 4 h after whole body irradiation (WBI) with 75 mGy X-rays was investigated with gel mobility shift assay. Results: Increased binding to CREB, NF-kB and APl consensus sequences was found with nuclear extracts prepared from thymocytes of irradiated versus sham-irradiated mice. Binding to the CREB, NF-kB and APl consensus sequences by nuclear extracts derived from irradiated mice was 6-fold, 4,3-fold and 2-fold higher than that from sham-irradiated respectively. The present report demonstrates that WBI with 75 mGy X-rays is capable of increasing expression of CREB, NF-kB and APl in murine thymocytes. competition with the cold oligonucleotide containing the consensus sequences for CREB and NF-kB resulted in loss of the shifted band, indicating specific binding. Conclusions: X-ray activation of select transcription factors, which bind certain consensus sites in promoters, may cause specific induction of gene transcription

  20. Trp53 activity is repressed in radio-adapted cultured murine limb bud cells

    Understanding the effects of ionizing radiation (IR) at low dose in fetal models is of great importance, because the fetus is considered to be at the most radiosensitive stage of the development and prenatal radiation might influence subsequent development. We previously demonstrated the existence of an adaptive response (AR) in murine fetuses after pre-exposure to low doses of X-rays. Trp53-dependent apoptosis was suggested to be responsible for the teratogenic effects of IR; decreased apoptosis was observed in adapted animals. In this study, in order to investigate the role of Trp53 in AR, we developed a new model of irradiated micromass culture of fetal limb bud cells, which replicated proliferation, differentiation and response to IR in murine embryos. Murine fetuses were exposed to whole-body priming irradiation of 0.3 Gy or 0.5 Gy at embryonic day 11 (E11). Limb bud cells (collected from digital ray areas exhibiting radiation-induced apoptosis) were cultured and exposed to a challenging dose of 4 Gy at E12 equivalent. The levels of Trp53 protein and its phosphorylated form at Ser18 were investigated. Our results suggested that the induction of AR in mouse embryos was correlated with a repression of Trp53 activity. (author)

  1. Interplay of Murine Gammaherpesvirus 68 with NF-kappaB Signaling of the Host.

    Cieniewicz, Brandon; Santana, Alexis L; Minkah, Nana; Krug, Laurie T

    2016-01-01

    Herpesviruses establish a chronic infection in the host characterized by intervals of lytic replication, quiescent latency, and reactivation from latency. Murine gammaherpesvirus 68 (MHV68) naturally infects small rodents and has genetic and biologic parallels with the human gammaherpesviruses (gHVs), Kaposi's sarcoma-associated herpesvirus and Epstein-Barr virus. The murine gammaherpesvirus model pathogen system provides a platform to apply cutting-edge approaches to dissect the interplay of gammaherpesvirus and host determinants that enable colonization of the host, and that shape the latent or lytic fate of an infected cell. This knowledge is critical for the development of novel therapeutic interventions against the oncogenic gHVs. The nuclear factor kappa B (NF-κB) signaling pathway is well-known for its role in the promotion of inflammation and many aspects of B cell biology. Here, we review key aspects of the virus lifecycle in the host, with an emphasis on the route that the virus takes to gain access to the B cell latency reservoir. We highlight how the murine gammaherpesvirus requires components of the NF-κB signaling pathway to promote replication, latency establishment, and maintenance of latency. These studies emphasize the complexity of gammaherpesvirus interactions with NF-κB signaling components that direct innate and adaptive immune responses of the host. Importantly, multiple facets of NF-κB signaling have been identified that might be targeted to reduce the burden of gammaherpesvirus-associated diseases. PMID:27582728

  2. Cytoplasmic superoxide dismutase and catalase activity and resistance to radiation lethality in murine tumor cells

    Reduced species of molecular oxygen are produced by the interaction of ionizing radiation with aqueous solutions containing molecular oxygen. The enzymes catalase and superoxide dismutase (SOD) are thought to function in vivo as scavengers of metabolically produced peroxide and superoxide respectively. SOD has been shown to protect against the lethal effects of ionizing radiation in vitro and in vivo. The authors have investigated the relationship between the cytosolic SOD catalase content and the sensitivity to radiation lethality of a number of murine cell lines (402AX, EL-4, MB-2T3, MB-4, MEL, P-815, SAI, SP-2, and SV-3T3). K/sub i/(CN-) for murine Cu-Zn-SOD was determined to be 6.8 x 10-6 M. No cytosolic Mn-SOD activity was found in any of the cell lines studied. No correlation was found between the cytosolic Cu-Zn-SOD or cytosolic catalase activity and the resistance to radiation lethality or the murine cell lines studied

  3. Developmental stage-related expression and identification of murine erythroid differentiation-denucleation factor (MEDDF)

    2001-01-01

    Using MEDDF cDNA fragment in plasmid pBS-SK-MEDDF as template the coding sequence was cloned into pGEM-T-Easy plasmid by PCR method to delete non-coding sequence. After DNA sequencing it was confirmed that the clone sequence was correct, the coding region then was inserted into the vector pET-30a between BamH I and Hind III to construct eukaryotic expression vector. It was found that the specific protein was up to 40% of total bactorial proteins in certain high-expression E. coli. High titer of anti-sera was detected by inoculating New Zealand rabbits with purified MEDDF protein as an antigen. By using immunocytochemical staining it was demonstrated that the expression of MEDDF was exhibited in a developmental stage-specific manner, suggesting that MEDDF may play a certain role in the initiation of murine erythroid terminal differentiation and nuclear condensation. As for the expression of MEDDF appearing in granulocytes and megakaryocyter in murine bone marrow, it may indicate that there is an original relationship between the proteins and differentiation of murine myelogenous lineage.

  4. Murine protein H is comprised of 20 repeating units, 61 amino acids in length

    Kristensen, Torsten; Tack, B F

    1986-01-01

    with this cDNA probe. Ten positives were colony-purified, and the largest plasmid cDNA insert, MH8 (4.4 kb), was sequenced by the dideoxy chain termination method. MH8 contained the complete coding sequence for the precursor of murine complement protein factor H (3702 bp), 100 bp of 5'-untranslated sequence......A cDNA library constructed from size-selected (greater than 28 S) poly(A)+ RNA isolated from the livers of C57B10. WR mice was screened by using a 249-base-pair (bp) cDNA fragment encoding 83 amino acid residues of human protein H as a probe. Of 120,000 transformants screened, 30 hybridized......, 448 bp of 3'-untranslated sequence, and a polyadenylylated tail of undetermined length. Murine pre-protein H was deduced to consist of an 18-amino acid signal peptide and 1216 residues of H-protein sequence. Murine H was composed of 20 repetitive units, each about 61 amino acid residues in length...

  5. Exon-intron organization and sequence comparison of human and murine T11 (CD2) genes

    Genomic DNA clones containing the human and murine genes coding for the 50-kDa T11 (CD2) T-cell surface glycoprotein were characterized. The human T11 gene is ≅ 12 kilobases long and comprised of five exons. A leader exon (L) contains the 5'-untranslated region and most of the nucleotides defining the signal peptide [amino acids (aa) -24 to -5]. Two exons encode the extracellular segment; exon Ex1 is 321 base pairs (bp) long and codes for four residues of the leader peptide and aa 1-103 of the mature protein, and exon Ex2 is 231 bp long and encodes aa 104-180. Exon TM is 123 bp long and codes for the single transmembrane region of the molecule (aa 181-221). Exon C is a large 765-bp exon encoding virtually the entire cytoplasmic domain (aa 222-327) and the 3'-untranslated region. The murine region T11 gene has a similar organization with exon-intron boundaries essentially identical to the human gene. Substantial conservation of nucleotide sequences between species in both 5'- and 3'-gene flanking regions equivalent to that among homologous exons suggests that murine and human genes may be regulated in a similar fashion. The probable relationship of the individual T11 exons to functional and structural protein domains is discussed

  6. Secretion of N- and O-linked Glycoproteins from 4T1 Murine Mammary Carcinoma Cells.

    Phang, Wai-Mei; Tan, Aik-Aun; Gopinath, Subash C B; Hashim, Onn H; Kiew, Lik Voon; Chen, Yeng

    2016-01-01

    Breast cancer is one of the most common cancers that affect women globally and accounts for ~23% of all cancers diagnosed in women. Breast cancer is also one of the leading causes of death primarily due to late stage diagnoses and a lack of effective treatments. Therefore, discovering protein expression biomarkers is mandatory for early detection and thus, critical for successful therapy. Two-dimensional electrophoresis (2D-E) coupled with lectin-based analysis followed by mass spectrometry were applied to identify potential biomarkers in the secretions of a murine mammary carcinoma cell line. Comparisons of the protein profiles of the murine 4T1 mammary carcinoma cell line and a normal murine MM3MG mammary cell line indicated that cadherin-1 (CDH), collagenase 3 (MMP-13), Viral envelope protein G7e (VEP), Gag protein (GAG) and Hypothetical protein LOC433182 (LOC) were uniquely expressed by the 4T1 cells, and pigment epithelium-derived factor (PEDF) was exclusively secreted by the MM3MG cells. Further analysis by a lectin-based study revealed that aberrant O-glycosylated CDH, N-glycosylated MMP-13 and LOC were present in the 4T1 medium. These differentially expressed N- and O-linked glycoprotein candidates, which were identified by combining lectin-based analysis with 2D-E, could serve as potential diagnostic and prognostic markers for breast cancer. PMID:27226773

  7. Applying advanced imaging techniques to a murine model of orthotopic osteosarcoma

    Matthew Lawrence Broadhead

    2015-08-01

    Full Text Available IntroductionReliable animal models are required to evaluate novel treatments for osteosarcoma. In this study, the aim was to implement advanced imaging techniques in a murine model of orthotopic osteosarcoma to improve disease modeling and the assessment of primary and metastatic disease.Materials and methodsIntra-tibial injection of luciferase-tagged OPGR80 murine osteosarcoma cells was performed in Balb/c nude mice. Treatment agent (pigment epithelium-derived factor; PEDF was delivered to the peritoneal cavity. Primary tumors and metastases were evaluated by in vivo bioluminescent assays, micro-computed tomography, [18F]-Fluoride-PET and [18F]-FDG-PET. Results[18F]-Fluoride-PET was more sensitive than [18F]-FDG-PET for detecting early disease. Both [18F]-Fluoride-PET and [18F]-FDG-PET showed progressive disease in the model, with 4-fold and 2-fold increases in SUV (p<0.05 by the study endpoint, respectively. In vivo bioluminescent assay showed that systemically delivered PEDF inhibited growth of primary osteosarcoma.DiscussionApplication of [18F]-Fluoride-PET and [18F]-FDG-PET to an established murine model of orthotopic osteosarcoma has improved the assessment of disease. The use of targeted imaging should prove beneficial for the evaluation of new approaches to osteosarcoma therapy.

  8. Secretion of N- and O-linked Glycoproteins from 4T1 Murine Mammary Carcinoma Cells

    Phang, Wai-Mei; Tan, Aik-Aun; Gopinath, Subash C.B.; Hashim, Onn H.; Kiew, Lik Voon; Chen, Yeng

    2016-01-01

    Breast cancer is one of the most common cancers that affect women globally and accounts for ~23% of all cancers diagnosed in women. Breast cancer is also one of the leading causes of death primarily due to late stage diagnoses and a lack of effective treatments. Therefore, discovering protein expression biomarkers is mandatory for early detection and thus, critical for successful therapy. Two-dimensional electrophoresis (2D-E) coupled with lectin-based analysis followed by mass spectrometry were applied to identify potential biomarkers in the secretions of a murine mammary carcinoma cell line. Comparisons of the protein profiles of the murine 4T1 mammary carcinoma cell line and a normal murine MM3MG mammary cell line indicated that cadherin-1 (CDH), collagenase 3 (MMP-13), Viral envelope protein G7e (VEP), Gag protein (GAG) and Hypothetical protein LOC433182 (LOC) were uniquely expressed by the 4T1 cells, and pigment epithelium-derived factor (PEDF) was exclusively secreted by the MM3MG cells. Further analysis by a lectin-based study revealed that aberrant O-glycosylated CDH, N-glycosylated MMP-13 and LOC were present in the 4T1 medium. These differentially expressed N- and O-linked glycoprotein candidates, which were identified by combining lectin-based analysis with 2D-E, could serve as potential diagnostic and prognostic markers for breast cancer.

  9. Fatty acid extracts from Lucilia sericata larvae promote murine cutaneous wound healing by angiogenic activity

    Zhang Jianing

    2010-03-01

    Full Text Available Abstract Background fatty acids are considered to be effective components to promote wound healing and Lucilia sericata larvae are applied clinically to treat intractable wounds. We aimed to investigat the effect of fatty acid extracts from dried Lucilia sericata larvae on murine cutaneuous wound healing as well as angiogenesis. Results On day 7 and 10 after murine acute excision wounds creation, the percent wound contraction of fatty acid extracts group was higher than that of vaseline group. On day 3, 7 and 10 after wounds creation, the wound healing quality of fatty acid extracts group was better than that of vaseline group on terms of granulation formation and collagen organization. On day 3 after wounds creation, the micro vessel density and vascular endothelial growth factor expression of fatty acid extracts group were higher than that of vaseline group. Component analysis of the fatty acid extracts by gas chromatography-mass spectrometry showed there were 10 kinds of fatty acids in total and the ratio of saturated fatty acid, monounsaturated fatty acid and polyunsaturated fatty acid (PUFA was: 20.57%:60.32%:19.11%. Conclusions Fatty acid extracts from dried Lucilia sericata larvae, four fifths of which are unsaturated fatty acids, can promote murine cutaneous wound healing probably resulting from the powerful angiogenic activity of the extracts.

  10. A rapid bioassay to monitor murine leukemia virus infection in mice using cellular gp71 binding

    A rapid and sensitive bioassay based on the availability of cell surface receptors for the binding of purified envelope glycoprotein, gp71, of Rauscher murine leukemia virus (R-MuLV) was developed to serially monitor viral-induced leukemogenesis in individual BALB/cAnN mice. The specificity of the bioassay was demonstrated by the competition of [125I]gp71 cellular binding with murine ecotropic viruses, purified unlabelled R-MuLV envelope glycoprotein and by antiserum to R-MuLV gp71. In contrast, there was no effect on the [125I]gp71 binding level with the addition of murine xenotropic viruses, R-MuLV p30, or several other proteins. The [125I]gp71 binding level of circulating leukocytes was significantly (P < 0.05) reduced in mice after R-MuLV infection. The reduction of cellular gp71 binding developed in two stages and the latter stage was highly dependent (P < 0.05) on circulating infections virus titer. Using this technique, the gp71 cellular binding levels of 48-60 individual mice can be assayed in a 4 h period. The advantages of this bioassay compared to standard immunological and tissue culture techniques used in studying retrovirus expression and viral-cell interactions are discussed. (Auth.)

  11. Antioxidative effects in vivo and colonization of Lactobacillus plantarum MA2 in the murine intestinal tract.

    Tang, Wei; Xing, Zhuqing; Hu, Wei; Li, Chao; Wang, Jinju; Wang, Yanping

    2016-08-01

    Lactobacillus plantarum MA2 was isolated from traditional Chinese Tibet kefir grains, which possess several excellent properties and functions. We previously demonstrated the antioxidant activities of this bacterium in vitro. However, the maintenance and survival of L. plantarum MA2 inside the murine intestinal tract, where it exerts its probiotic properties, and whether its effects are elicited directly on the host remain unknown. Therefore, this study investigated the mechanisms of L. plantarum MA2 in aging mice following D-galactose administration. The levels of malondialdehyde decreased significantly in the L. plantarum MA2 groups after oral ingestion compared to the D-galactose model group, and total antioxidant capacity and glutathione peroxidase and superoxide dismutase activities increased significantly in the serum and liver. We combined fluorescein isothiocyanate labeling and green fluorescent protein expression to dynamically monitor the colonization and distribution of L. plantarum MA2 in the murine intestinal tract. The results indicated that L. plantarum MA2 was detected in the ileum, colon, and feces after single and continuous oral administration at day 21 and was maintained at 10(4)-10(5) CFU/g. These results suggest that L. plantarum MA2 colonizes and survives in the murine intestinal tract to exert its antioxidative effects. PMID:27178180

  12. Bicarbonate Secretion in the Murine Gallbladder - Lessons for the Treatment of Cystic Fibrosis

    Cuthbert AW

    2001-07-01

    Full Text Available The epithelium lining the gallbladder of mammalian species has absorptive and secretory functions. An important function is the secretion of a bicarbonate rich fluid that helps neutralise stomach acid and provides an appropriate environment for intestinal enzymes. In cystic fibrosis (CF this secretory function is lost. This study concerns the bicarbonate secreting activity of murine gallbladders in vitro using wild type and CF mice and four main questions are considered as follows: a Does the murine gallbladder secrete bicarbonate electrogenically and is this prevented in CF? b Can the secretory activity in CF gallbladders be restored by gene therapy or pharmacologically? c How is the cystic fibrosis transmembrane conductance regulator (CFTR involved in bicarbonate secretion? d Does the data offer prospects for the treatment of CF?. Work from both the author's laboratory and the literature will be reviewed. Consideration of the currently available data indicates that the wild type murine gallbladder does secrete bicarbonate electrogenically and that this is absent in CF mice. Further it has been demonstrated that bicarbonate secretory activity can be restored by both gene therapy and by the use of drugs. The role of CFTR in bicarbonate secretion remains equivocal. Much evidence suggests that CFTR can act as a channel for HCO(3(- ions as well as Cl(- ions, while others propose a parallel arrangement of CFTR with a Cl(-/HCO(3(- exchanger is necessary. The matter is further complicated by the regulatory role of CFTR on other transporting activities. Opportunities for possible application to man are discussed.

  13. Puerarin Facilitates T-Tubule Development of Murine Embryonic Stem Cell-Derived Cardiomyocytes

    Lu Wang

    2014-07-01

    Full Text Available Aims: The embryonic stem cell-derived cardiomyocytes (ES-CM is one of the promising cell sources for repopulation of damaged myocardium. However, ES-CMs present immature structure, which impairs their integration with host tissue and functional regeneration. This study used murine ES-CMs as an in vitro model of cardiomyogenesis to elucidate the effect of puerarin, the main compound found in the traditional Chinese medicine the herb Radix puerariae, on t-tubule development of murine ES-CMs. Methods: Electron microscope was employed to examine the ultrastructure. The investigation of transverse-tubules (t-tubules was performed by Di-8-ANEPPS staining. Quantitative real-time PCR was utilized to study the transcript level of genes related to t-tubule development. Results: We found that long-term application of puerarin throughout cardiac differentiation improved myofibril array and sarcomeres formation, and significantly facilitated t-tubules development of ES-CMs. The transcript levels of caveolin-3, amphiphysin-2 and junctophinlin-2, which are crucial for the formation and development of t-tubules, were significantly upregulated by puerarin treatment. Furthermore, puerarin repressed the expression of miR-22, which targets to caveolin-3. Conclusion: Our data showed that puerarin facilitates t-tubule development of murine ES-CMs. This might be related to the repression of miR-22 by puerarin and upregulation of Cav3, Bin1 and JP2 transcripts.

  14. Notch pathway inhibition controls myeloma bone disease in the murine MOPC315.BM model

    Despite evidence that deregulated Notch signalling is a master regulator of multiple myeloma (MM) pathogenesis, its contribution to myeloma bone disease remains to be resolved. Notch promotes survival of human MM cells and triggers human osteoclast activity in vitro. Here, we show that inhibition of Notch through the γ-secretase inhibitor XII (GSI XII) induces apoptosis of murine MOPC315.BM myeloma cells with high Notch activity. GSI XII impairs murine osteoclast differentiation of receptor activator of NF-κB ligand (RANKL)-stimulated RAW264.7 cells in vitro. In the murine MOPC315.BM myeloma model GSI XII has potent anti-MM activity and reduces osteolytic lesions as evidenced by diminished myeloma-specific monoclonal immunoglobulin (Ig)-A serum levels and quantitative assessment of bone structure changes via high-resolution microcomputed tomography scans. Thus, we suggest that Notch inhibition through GSI XII controls myeloma bone disease mainly by targeting Notch in MM cells and possibly in osteoclasts in their microenvironment. We conclude that Notch inhibition is a valid therapeutic strategy in MM

  15. Spontaneous mutations in the CsrRS two-component regulatory system of Streptococcus pyogenes result in enhanced virulence in a murine model of skin and soft tissue infection.

    Engleberg, N C; Heath, A; Miller, A; Rivera, C; DiRita, V J

    2001-04-01

    CsrS/CsrR is a 2-component system in Streptococcus pyogenes that negatively regulates hyaluronic capsule and several exotoxins. To detect spontaneous mutations in csrRS, mucoid and large colony variants of M1 strain MGAS166 were isolated from experimental murine skin infections. By use of complementation with a csrRS(+) plasmid, relevant mutations were also detected in 7 of 12 human clinical isolates. The presence of spontaneous mutants in mouse infection was associated with larger, more necrotic lesions. Most spontaneous changes in CsrR resulted from single amino acid substitutions, whereas most csrS mutations were frameshift or nonsense mutations. In 2 instances, IS1548 insertions were found in csrS. Experimental inoculation of mixtures of wild-type (wt) and csrRS(-) bacteria yielded larger, more necrotic lesions than did either strain at twice the inoculum, which suggests that these variants may exhibit pathogenic synergy. Spontaneous emergence of csrRS(-) mutants in vivo enhances the virulence of wt bacteria and increases severity of murine skin infection. PMID:11237829

  16. Alport alloantibodies but not Goodpasture autoantibodies induce murine glomerulonephritis: protection by quinary crosslinks locking cryptic α3(IV) collagen autoepitopes in vivo.

    Luo, Wentian; Wang, Xu-Ping; Kashtan, Clifford E; Borza, Dorin-Bogdan

    2010-09-15

    The noncollagenous (NC1) domains of alpha3alpha4alpha5(IV) collagen in the glomerular basement membrane (GBM) are targets of Goodpasture autoantibodies or Alport posttransplant nephritis alloantibodies mediating rapidly progressive glomerulonephritis. Because the autoepitopes but not the alloepitopes become cryptic upon assembly of alpha3alpha4alpha5NC1 hexamers, we investigated how the accessibility of B cell epitopes in vivo influences the development of glomerulonephritis in mice passively immunized with human anti-GBM Abs. Alport alloantibodies, which bound to native murine alpha3alpha4alpha5NC1 hexamers in vitro, deposited linearly along the mouse GBM in vivo, eliciting crescentic glomerulonephritis in Fcgr2b(-/-) mice susceptible to Ab-mediated inflammation. Goodpasture autoantibodies, which bound to murine alpha3NC1 monomer and dimer subunits but not to native alpha3alpha4alpha5NC1 hexamers in vitro, neither bound to the mouse GBM in vivo nor induced experimental glomerulonephritis. This was due to quinary NC1 crosslinks, recently identified as sulfilimine bonds, which comprehensively locked the cryptic Goodpasture autoepitopes in the mouse GBM. In contrast, non-crosslinked alpha3NC1 subunits were identified as a native target of Goodpasture autoantibodies in the GBM of squirrel monkeys, a species susceptible to Goodpasture autoantibody-mediated nephritis. Thus, crypticity of B cell autoepitopes in tissues uncouples potentially pathogenic autoantibodies from autoimmune disease. Crosslinking of alpha3alpha4alpha5NC1 hexamers represents a novel mechanism averting autoantibody binding and subsequent tissue injury by posttranslational modifications of an autoantigen. PMID:20709951

  17. Alport alloantibodies but not Goodpasture autoantibodies induce murine glomerulonephritis: Protection by quinary crosslinks locking cryptic α3(IV) collagen autoepitopes in vivo 1

    Luo, Wentian; Wang, Xu-Ping; Kashtan, Clifford E.; Borza, Dorin-Bogdan

    2010-01-01

    The noncollagenous (NC1) domains of α3α4α5(IV) collagen in the glomerular basement membrane (GBM) are targets of Goodpasture autoantibodies or Alport post-transplant nephritis alloantibodies mediating rapidly progressive glomerulonephritis. Because the autoepitopes but not the alloepitopes become cryptic upon assembly of α3α4α5NC1 hexamers, we investigated how the accessibility of B cell epitopes in vivo influences the development of glomerulonephritis in mice passively immunized with human anti-GBM antibodies. Alport alloantibodies, which bound to native murine α3α4α5NC1 hexamers in vitro, deposited linearly along the mouse GBM in vivo, eliciting crescentic glomerulonephritis in Fcgr2b−/− mice susceptible to antibody-mediated inflammation. Goodpasture autoantibodies, which bound to murine α3NC1 monomer and dimer subunits but not to native α3α4α5NC1 hexamers in vitro, neither bound to the mouse GBM in vivo nor induced experimental glomerulonephritis. This was due to quinary NC1 cross-links, recently identified as sulfilimine bonds, which comprehensively locked the cryptic Goodpasture autoepitopes in the mouse GBM. In contrast, non-crosslinked α3NC1 subunits were identified as a native target of Goodpasture autoantibodies in the GBM of squirrel monkeys—a species susceptible to Goodpasture autoantibody-mediated nephritis. Thus, crypticity of B cell autoepitopes in tissues uncouples potentially pathogenic autoantibodies from autoimmune disease. Crosslinking of α3α4α5NC1 hexamers represents a novel mechanism averting autoantibody binding and subsequent tissue injury by post-translational modifications of an autoantigen. PMID:20709951

  18. Investigation of anti-WI-1 adhesin antibody-mediated protection in experimental pulmonary blastomycosis.

    Wüthrich, M; Klein, B S

    2000-05-01

    Infection with Blastomyces dermatitidis elicits strong antibody responses to the surface adhesin WI-1. The antibodies are directed chiefly against the adhesive domain, a 25-amino-acid repeat. Tandem-repeat-specific monoclonal antibodies (mAbs) were studied for their opsonic activity in vitro and their capacity to adoptively transfer protection in murine experimental blastomycosis. mAbs to WI-1 enhanced binding and entry of B. dermatitidis yeasts into J774. 16 cells but did not enhance killing or growth inhibition of the yeast. Passive transfer of 8 mAbs to WI-1 into 3 different inbred strains of mice also did not improve the course of experimental infection and sometimes worsened it. mu-deficient mice were more resistant to experimental blastomycosis than were intact littermates, and passive transfer of the mAbs into these mice did not protect them against experimental infection. Thus, antibody to WI-1 does not appear to improve the outcome of murine blastomycosis and may enhance the infection. PMID:10823774

  19. The Involvement of Heat Shock Proteins in Murine Liver Regeneration

    Qing Shi; Zhongjun Dong; Haiming Wei

    2007-01-01

    Partial hepatectomy (PHx) in mammals is a very common experimental model to investigate the process of liver regeneration. The surgery itself could give birth to a series of stresses, such as the temporary raise of body temperature and the ischaemia-reperfusion injury. Heat shock proteins (HSPs) were a family of stress-inducible proteins involved in maintaining cell homeostasis and regulating the immune system. In our study, we intended to investigate the expression and role of HSPs in liver regeneration. Using RT-PCR and Western blotting, we determined the expression in regenerating liver of HSP27, HSP60, HSP70 and HSP90 in mRNA level and protein level, respectively, with mice treated with sham operation as controls. We also used quercertin as an inhibitior of HSPs to explore their effects on liver regeneration. We found that hepatic expression of HSPs increased at the early phase of liver regeneration and declined to the constitutively low level later. Moreover, quercetin pretreatment delayed the progress of liver regeneration in mice via inhibition of HSPs. The results indicated that HSPs played an important role in liver regeneration.

  20. A Proteomic Investigation of Hepatic Resistance to Ascaris in a Murine Model.

    Deslyper, Gwendoline; Colgan, Thomas J; Cooper, Andrew J R; Holland, Celia V; Carolan, James C

    2016-08-01

    The helminth Ascaris causes ascariasis in both humans and pigs. Humans, especially children, experience significant morbidity including respiratory complications, growth deficits and intestinal obstruction. Given that 800 million people worldwide are infected by Ascaris, this represents a significant global public health concern. The severity of the symptoms and associated morbidity are related to the parasite burden and not all hosts are infected equally. While the pathology of the disease has been extensively examined, our understanding of the molecular mechanisms underlying resistance and susceptibility to this nematode infection is poor. In order to investigate host differences associated with heavy and light parasite burden, an experimental murine model was developed utilising Ascaris-susceptible and -resistant mice strains, C57BL/6J and CBA/Ca, respectively, which experience differential burdens of migratory Ascaris larvae in the host lungs. Previous studies identified the liver as the site where this difference in susceptibility occurs. Using a label free quantitative proteomic approach, we analysed the hepatic proteomes of day four post infection C57BL/6J and CBA/Ca mice with and without Ascaris infection to identify proteins changes potentially linked to both resistance and susceptibility amongst the two strains, respectively. Over 3000 proteins were identified in total and clear intrinsic differences were elucidated between the two strains. These included a higher abundance of mitochondrial proteins, particularly those associated with the oxidative phosphorylation pathway and reactive oxygen species (ROS) production in the relatively resistant CBA/Ca mice. We hypothesise that the increased ROS levels associated with higher levels of mitochondrial activity results in a highly oxidative cellular environment that has a dramatic effect on the nematode's ability to successfully sustain a parasitic association with its resistant host. Under infection, both