ABSTRACT
Mosquitoes collected in northern Italy were screened for flavivirus RNA. Positive amplicons were sequenced and found most similar to insect flavivirus (ISF), Usutu virus (USUV) and surprisingly also to Japanese encephalitis virus (JEV). The sequence (167 bp), obtained from one pool of Culex pipiens, was found identical to JEV strains from bats in China. Unfortunately additional sequence data or virus isolations were not obtained in this study. Confirmation of potential introduction of JEV to Italy and other European countries is urgently needed.
Subject(s)
Culex/virology , Encephalitis Virus, Japanese/isolation & purification , Insect Vectors/virology , RNA, Viral/genetics , Amplified Fragment Length Polymorphism Analysis , Animals , China , Chiroptera/virology , Culex/genetics , Encephalitis Virus, Japanese/genetics , Italy , Sequence AnalysisABSTRACT
PURPOSE: We propose herein labeling protocols for multimodal in vivo visualization of human skeletal muscle cells (HSkMCs) by MRI and BLI to investigate the survival, localization, and proliferation/differentiation of these cells in cell-mediated therapy. PROCEDURES: HSkMCs were labeled with different quantities of Endorem® and transfection agents or infected with lentiviral vector expressing the luciferase gene under the myogenin promoter. Cells were evaluated before and after intra-arterial injection in NUDE mice with N2-induced muscle inflammation. RESULTS: Neither iron labeling nor infection affected cell features; the number of iron-positive cells increased proportionally to the iron content in the medium and in the presence of transfection agents. Loaded cells were detected for up to 1 month by MRI and 2 months by BLI. CONCLUSIONS: These protocols could be used to visualize new stem cells, in vivo and over time, in preclinical studies of cell-based treatments for myopathies of different etiologies.
Subject(s)
Cell Tracking/methods , Luminescent Measurements/methods , Magnetic Resonance Imaging/methods , Magnetite Nanoparticles/chemistry , Muscle Fibers, Skeletal/cytology , Analysis of Variance , Animals , Cell Differentiation/drug effects , Cell Survival/drug effects , Cells, Cultured , Endocytosis/drug effects , Female , Histocytochemistry , Humans , Magnetite Nanoparticles/adverse effects , Mice , Mice, Nude , Muscle Fibers, Skeletal/chemistry , Staining and Labeling , Tissue Distribution , TransfectionABSTRACT
In beta-thalassemia, the mechanism driving ineffective erythropoiesis (IE) is insufficiently understood. We analyzed mice affected by beta-thalassemia and observed, unexpectedly, a relatively small increase in apoptosis of their erythroid cells compared with healthy mice. Therefore, we sought to determine whether IE could also be characterized by limited erythroid cell differentiation. In thalassemic mice, we observed that a greater than normal percentage of erythroid cells was in S-phase, exhibiting an erythroblast-like morphology. Thalassemic cells were associated with expression of cell cycle-promoting genes such as EpoR, Jak2, Cyclin-A, Cdk2, and Ki-67 and the antiapoptotic protein Bcl-X(L). The cells also differentiated less than normal erythroid ones in vitro. To investigate whether Jak2 could be responsible for the limited cell differentiation, we administered a Jak2 inhibitor, TG101209, to healthy and thalassemic mice. Exposure to TG101209 dramatically decreased the spleen size but also affected anemia. Although our data do not exclude a role for apoptosis in IE, we propose that expansion of the erythroid pool followed by limited cell differentiation exacerbates IE in thalassemia. In addition, these results suggest that use of Jak2 inhibitors has the potential to profoundly change the management of this disorder.
