Search results for "fluorescent"

Pippin protein expression changes during cell differentiation

2008

PIPPin is a CSD-containing protein with the ability to interact both with mRNAs encoding histone variants and chromatin. A major fraction of chromatin-bound PIPPin is sumoylated and sumoylation seems to be controlled by thyroid hormones, both in vivo and in vitro. We studied its expression in different tissues and cell lines and even in tumor cells and found that, even if more expressed in the brain respect to other tissues of the adult rat, it is also expressed in brain tumors and in cell lines as different as kidney NRK cells and PC12. The expression of the protein is strongly increased by treatments that induce differentiation, such as treatment of PC12 with NGF. We also found an increas…

SMOOTHLY SHIFTING FLUORESCENT WINDOW: TUNABLE “OFF-ON-OFF” MICELLAR BIOSENSORS FOR pH

2009

An in vitro procedure for evaluation of early stage oxidative stress in an established fish cell line applied to investigation of PHAH and pesticide …

2004

Oxidative stress by increased production of reactive oxygen species such as superoxide has been implicated in the toxicity of PCB's and non-target toxicity of many pesticides. We report the development of a microplate-based method for determination of early stage oxidative stress using an established cell line (EPC) from a skin tumour of carp Cyprinus carpio L. and 2',7'-dichlorodihydrofluorescein diacetate (H(2)-DCFDA) as a fluorescent probe for detection of reactive oxygen species (ROS) formation. Sublethal concentrations of the herbicide Paraquat, an established redox cycling agent and a crude PCB mixture, Arochlor 1254 elicited a linear increase in ROS formation over 2 h exposure which …

Expression and subcellular targeting of canine parvovirus capsid proteins in baculovirus-transduced NLFK cells

2004

AbstractA mammalian baculovirus delivery system was developed to study targeting in Norden Laboratories feline kidney (NLFK) cells of the capsid proteins of canine parvovirus (CPV), VP1 and VP2, or corresponding counterparts fused to EGFP. VP1 and VP2, when expressed alone, both had equal nuclear and cytoplasmic distribution. However, assembled form of VP2 had a predominantly cytoplasmic localization. When VP1 and VP2 were simultaneously present in cells, their nuclear localization increased. Thus, confocal immunofluorescence analysis of cells transduced with the different baculovirus constructs or combinations thereof in the absence or presence of infecting CPV revealed that the VP1 protei…

Assembly of fluorescent chimeric virus-like particles of canine parvovirus in insect cells

2004

Canine parvovirus (CPV) is a small non-enveloped ssDNA virus composed of the viral proteins VP1, VP2, and VP3 with a T=1 icosahedral symmetry. VP2 is nested in VP1 and the two proteins are produced by differential splicing of a primary transcript of the right ORF of the viral genome. The VP2 protein can be further proteolytically cleaved to form VP3. Previous studies have shown that VP1 and VP3 are unnecessary for capsid formation and consequently, that VP2 alone is sufficient for assembly. We have hypothesized that insertion of the enhanced green fluorescent protein (EGFP) at the N-terminus of VP2 could be carried out without altering assembly. To investigate the possibility to develop flu…

Detection of canine parvovirus antigens with antibodies to synthetic peptides

1996

Antibodies produced in rabbits against an 18-amino acid peptide (peptide 1, NSLPQSEGATNFGDIGVP) of capsid protein VP2/residues 292-309 of canine parvovirus (CPV) or against an 18-amino acid peptide (peptide 2, GKRNTVLFHGPASTKGKS) of nonstructural protein NS1/residues 391-409 of CPV identified, in immunofluorescence analysis, viral antigens in canine A 72 cells infected with CPV. Antibodies to peptide 2 also identified viral antigens in bovine cells infected with bovine parvovirus. In western blot analysis, antibodies to peptide 1 and peptide 2 also detected viral antigens derived from blue fox parvovirus, feline parvovirus, mink enteritis virus and raccoon dog parvovirus. The peptide antibo…

Parvovirus induced alterations in nuclear architecture and dynamics.

2009

The nucleus of interphase eukaryotic cell is a highly compartmentalized structure containing the three-dimensional network of chromatin and numerous proteinaceous subcompartments. DNA viruses induce profound changes in the intranuclear structures of their host cells. We are applying a combination of confocal imaging including photobleaching microscopy and computational methods to analyze the modifications of nuclear architecture and dynamics in parvovirus infected cells. Upon canine parvovirus infection, expansion of the viral replication compartment is accompanied by chromatin marginalization to the vicinity of the nuclear membrane. Dextran microinjection and fluorescence recovery after ph…

Synaptopodin regulates denervation-induced homeostatic synaptic plasticity

2013

Synaptopodin (SP) is a marker and essential component of the spine apparatus (SA), an enigmatic cellular organelle composed of stacked smooth endoplasmic reticulum that has been linked to synaptic plasticity. However, SP/SA-mediated synaptic plasticity remains incompletely understood. To study the role of SP/SA in homeostatic synaptic plasticity we here used denervation-induced synaptic scaling of mouse dentate granule cells as a model system. This form of plasticity is of considerable interest in the context of neurological diseases that are associated with the loss of neurons and subsequent denervation of connected brain regions. In entorhino-hippocampal slice cultures prepared from SP-de…

A subset of ventral tegmental area dopamine neurons responds to acute ethanol

2015

The mechanisms by which alcohol drinking promotes addiction in humans and self-administration in rodents remain obscure, but it is well known that alcohol can enhance dopamine (DA) neurotransmission from neurons of the ventral tegmental area (VTA) and increase DA levels within the nucleus accumbens and prefrontal cortex. We recorded from identified DA neuronal cell bodies within ventral midbrain slices prepared from a transgenic mouse line (TH-GFP) using long-term stable extracellular recordings in a variety of locations and carefully mapped the responses to applied ethanol (EtOH). We identified a subset of DA neurons in the medial VTA located within the rostral linear and interfascicular n…

“Super p53” Mice Display Retinal Astroglial Changes

2013

Tumour-suppressor genes, such as the p53 gene, produce proteins that inhibit cell division under adverse conditions, as in the case of DNA damage, radiation, hypoxia, or oxidative stress (OS). The p53 gene can arrest proliferation and trigger death by apoptosis subsequent to several factors. In astrocytes, p53 promotes cell-cycle arrest and is involved in oxidative stress-mediated astrocyte cell death. Increasingly, astrocytic p53 is proving fundamental in orchestrating neurodegenerative disease pathogenesis. In terms of ocular disease, p53 may play a role in hypoxia due to ischaemia and may be involved in the retinal response to oxidative stress (OS). We studied the influence of the p53 ge…