Home About us Contact
|
Home About us Contact | ||
|
|
||
Used Mice Deficient (used + mouse_deficient)
Selected AbstractsDetrimental role of endogenous nitric oxide in host defence against Sporothrix schenckiiIMMUNOLOGY, Issue 4 2008Karla Simone S. Fernandes Summary We earlier demonstrated that nitric oxide (NO) is a fungicidal molecule against Sporothrix schenckii in vitro. In the present study we used mice deficient in inducible nitric oxide synthase (iNOS,/,) and C57BL/6 wild-type (WT) mice treated with N,-nitro-arginine (Nitro-Arg-treated mice), an NOS inhibitor, both defective in the production of reactive nitrogen intermediates, to investigate the role of endogenous NO during systemic sporotrichosis. When inoculated with yeast cells of S. schenckii, WT mice presented T-cell suppression and high tissue fungal dissemination, succumbing to infection. Furthermore, susceptibility of mice seems to be related to apoptosis and high interleukin-10 and tumour necrosis factor-, production by spleen cells. In addition, fungicidal activity and NO production by interferon-, (IFN-,) and lipopolysaccharide-activated macrophages from WT mice were abolished after fungal infection. Strikingly, iNOS,/, and Nitro-Arg-treated mice presented fungal resistance, controlling fungal load in tissues and restoring T-cell activity, as well as producing high amounts of IFN-, Interestingly, macrophages from these groups of mice presented fungicidal activity after in vitro stimulation with higher doses of IFN-,. Herein, these results suggest that although NO was an essential mediator to the in vitro killing of S. schenckii by macrophages, the activation of NO system in vivo contributes to the immunosuppression and cytokine balance during early phases of infection with S. schenckii. [source] Identifying the Relative Contributions of Rac1 and Rac2 to Osteoclastogenesis,,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 2 2008Yongqiang Wang Abstract Rac small GTPases may play an important regulatory role in osteoclastogenesis. Our in vitro and in vivo results show that both Rac1 and Rac2 are required for optimal osteoclast differentiation, but Rac1 is more critical. Rac1 is the key Rac isoform responsible for regulating ROS generation and the actin cytoskeleton during the multiple stages of osteoclast differentiation. Introduction: Recent evidence suggests that the Rac small GTPases may play an important regulatory role in osteoclastogenesis. This finding is important because bisphosphonates may regulate their antiresorptive/antiosteoclast effects through the modification of Rho family of small GTPases. Materials and Methods: To elucidate the specific roles of the Rac1 and Rac2 isoforms during osteoclastogenesis, we used mice deficient in Rac1, Rac2, or both Rac1 and Rac2 in monocyte/osteoclast precursors. Macrophage-colony stimulating factor (M-CSF), and RANKL-mediated osteoclastogenesis in vitro was studied by using bone marrow-derived mononucleated preosteoclast precursors (MOPs). The expression of osteoclast-specific markers was examined using quantitative real-time PCR and Western blot analysis. Free actin barbed ends in bone marrow MOPs after M-CSF stimulation was determined. The ability of MOPs to migrate toward M-CSF was assayed using Boyden chambers. Margin spreading on heparin sulfate-coated glass and RANKL-induced reactive oxygen species generation were also performed. Functional assays of in vitro-generated osteoclasts were ascertained using dentine sections from narwal tusks. Osteoclast levels in vivo were counted in TRACP and immunohistochemically stained distal tibial sections. In vivo microarchitexture of lumbar vertebrate was examined using ,CT 3D imaging and analysis. Results: We show here that, although both Rac isoforms are required for normal osteoclast differentiation, Rac1 deletion results in a more profound reduction in osteoclast formation in vitro because of its regulatory role in pre-osteoclast M-CSF-mediated chemotaxis and actin assembly and RANKL-mediated reactive oxygen species generation. This Rac1 cellular defect also manifests at the tissue level with increased trabecular bone volume and trabeculae number compared with wildtype and Rac2-null mice. This unique mouse model has shown for the first time that Rac1 and Rac2 play different and nonoverlapping roles during osteoclastogenesis and will be useful for identifying the key roles played by these two proteins during the multiple stages of osteoclast differentiation. Conclusions: Rac1 and Rac2 play different and nonoverlapping roles during osteoclastogenesis. This model showed that Rac1 is the key Rac isoform responsible for regulating ROS generation and the actin cytoskeleton during the multiple stages of osteoclast differentiation. [source] Inactivation of Pten in Osteo-Chondroprogenitor Cells Leads to Epiphyseal Growth Plate Abnormalities and Skeletal Overgrowth,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 8 2007Alice Fiona Ford-Hutchinson Abstract To study the role of the Pten tumor suppressor in skeletogenesis, we generated mice lacking this key phosphatidylinositol 3,-kinase pathway regulator in their osteo-chondroprogenitors. A phenotype of growth plate dysfunction and skeletal overgrowth was observed. Introduction: Skeletogenesis is a complex process relying on a variety of ligands that activate a range of intracellular signal transduction pathways. Although many of these stimuli are known to activate phosphatidylinositol 3,-kinase (PI3K), the function of this pathway during cartilage development remains nebulous. To study the role of PI3K during skeletogenesis, we used mice deficient in a negative regulator of PI3K signaling, the tumor suppressor, Pten. Materials and Methods:Pten gene deletion in osteo-chondrodroprogenitors was obtained by interbreeding mice with loxP-flanked Pten exons with mice expressing the Cre recombinase under the control of the type II collagen gene promoter (Ptenflox/flox:Col2a1Cre mice). Phenotypic analyses included microcomputed tomography and immunohistochemistry techniques. Results: ,CT revealed that Ptenflox/flox:Col2a1Cre mice exhibited both increased skeletal size, particularly of vertebrae, and massive trabeculation accompanied by increased cortical thickness. Primary spongiosa development and perichondrial bone collar formation were prominent in Ptenflox/flox:Col2a1Cre mice, and long bone growth plates were disorganized and showed both matrix overproduction and evidence of accelerated hypertrophic differentiation (indicated by an altered pattern of type X collagen and alkaline phosphatase expression). Consistent with increased PI3K signaling, Pten-deficient chondrocytes showed increased phospho-PKB/Akt and phospho-S6 immunostaining, reflective of increased mTOR and PDK1 activity. Interestingly, no significant change in growth plate proliferation was seen in Pten-deficient mice, and growth plate fusion was found at 6 months. Conclusions: By virtue of its ability to modulate a key signal transduction pathway responsible for integrating multiple stimuli, Pten represents an important regulator of both skeletal size and bone architecture. [source] RGS4 Controls Renal Blood Flow and Inhibits Cyclosporine-Mediated NephrotoxicityAMERICAN JOURNAL OF TRANSPLANTATION, Issue 2 2010A. Siedlecki Calcineurin inhibitors (CNI) are powerful immunomodulatory agents that produce marked renal dysfunction due in part to endothelin-1-mediated reductions in renal blood flow. Ligand-stimulated Gq protein signaling promotes the contraction of smooth muscle cells via phospholipase C,-mediated stimulation of cytosolic calcium release. RGS4 is a GTPase activating protein that promotes the deactivation of Gq and Gi family members. To investigate the role of G protein-mediated signaling in the pathogenesis of CNI-mediated renal injury, we used mice deficient for RGS4 (rgs4,/,). Compared to congenic wild type control animals, rgs4,/, mice were intolerant of the CNI, cyclosporine (CyA), rapidly developing fatal renal failure. Rgs4,/, mice exhibited markedly reduced renal blood flow after CyA treatment when compared to congenic wild type control mice as measured by magnetic resonance imaging (MRI). Hypoperfusion was reversed by coadministration of CyA with the endothelin antagonist, bosentan. The MAPK/ERK pathway was activated by cyclosporine administration and was inhibited by cotreatment with bosentan. These results show that endothelin-1-mediated Gq protein signaling plays a key role in the pathogenesis of vasoconstrictive renal injury and that RGS4 antagonizes the deleterious effects of excess endothelin receptor activation in the kidney. [source] | ||||||||||