Analysis of Green Fluorescent Protein Expression in Transgenic Rats for Tracking Transplanted Neural Stem/Progenitor Cells

Mothe, Andrea J.; Kulbatski, Iris; Bendegem, Rita L. van; Lee, Linda; Kobayashi, Eiji; Keating, Armand; Tator, Charles H.

doi:10.1369/jhc.5a6639.2005

Cited by 45 publications

(24 citation statements)

References 34 publications

(57 reference statements)

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“…Surprisingly, and in apparent contradiction with the in vitro results, no GFP expression was detected in astrocytes or oligodendrocytes, suggesting that the PGK promoter is inserted in a locus under epigenetic control resulting in either activation of expression by the in vitro culture conditions or in suppression of its expression in vivo. Thus, neural stem/progenitor cells derived from this GFP high -transgenic rats may be very useful for neuronal replacement strategies, unlike those obtained from existing lines (Mothe et al 2005;Francis et al 2007) rather interesting for oligodendrocytes replacement.…”

Section: Discussionmentioning

confidence: 99%

New lines of GFP transgenic rats relevant for regenerative medicine and gene therapy

et al. 2010

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Adoptive cell transfer studies in regenerative research and identification of genetically modified cells after gene therapy in vivo require unequivocally identifying and tracking the donor cells in the host tissues, ideally over several days or for up to several months. The use of reporter genes allows identifying the transferred cells but unfortunately most are immunogenic to wild-type hosts and thus trigger rejection in few days. The availability of transgenic animals from the same strain that would express either high levels of the transgene to identify the cells or low levels but that would be tolerant to the transgene would allow performing long-term analysis of labelled cells. Herein, using lentiviral vectors we develop two new lines of GFP-expressing transgenic rats displaying different levels and patterns of GFP-expression. The "high-expresser" line (GFP(high)) displayed high expression in most tissues, including adult neurons and neural precursors, mesenchymal stem cells and in all leukocytes subtypes analysed, including myeloid and plasmacytoid dendritic cells, cells that have not or only poorly characterized in previous GFP-transgenic rats. These GFP(high)-transgenic rats could be useful for transplantation and immunological studies using GFP-positive cells/tissue. The "low-expresser" line expressed very low levels of GFP only in the liver and in less than 5% of lymphoid cells. We demonstrate these animals did not develop detectable humoral and cellular immune responses against both transferred GFP-positive splenocytes and lentivirus-mediated GFP gene transfer. Thus, these GFP-transgenic rats represent useful tools for regenerative medicine and gene therapy.

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Section: Discussionmentioning

confidence: 99%

New lines of GFP transgenic rats relevant for regenerative medicine and gene therapy

et al. 2010

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“…Many techniques were used and compared to optimize the immunohistochemical detection of GFP using conventional [34] and confocal microscope [35] even in paraffin embedded tissue As more and more studies relying on GFP appeared there were hints that the transgenes might be silenced depending on age, tissue, differentiation and many other factors. Mothe et al [36] compared detection of GFP in transgenic mice and rats and found that sometimes GFP has to be immunostained to be seen and suggested the possibility of a partial silencing of the transgene after cell transplantation. This could potentially occur because of genetic changes in the DNA or alterations in transcription or translation.…”

Section: Discussionmentioning

confidence: 99%

Sensitive detection of GFP utilizing tyramide signal amplification to overcome gene silencing

Tóth

Shahar

Leker

et al. 2007

Experimental Cell Research

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The green fluorescent protein (GFP) is among the most commonly used expression markers in biology. GFP-tagged cells have played a particularly important role in studies of cell lineage. Sensitive detection of GFP is crucially important for such studies to be successful, and problems with detection may account for discrepancies in the literature regarding the possible fate choices of stem cells. Here we describe a very sensitive technique for visualization of GFP. Using it we can detect about 90% of cells of donor origin while we could only see about 50% of these cells when we employ the methods that are in general use in other laboratories. In addition, we provide evidence that some cells permanently silence GFP expression. In the case of the progeny of bone marrow stem cells, it appears that the more distantly related they are to their precursors, the more likely it is that they will turn off the lineage marker.

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“…Here we show that coculture of neurosphere-derived NSCs with MSCs had a profound effect on the differentiation of the neural cells. To unambiguously distinguish neural cells from MSCs the neurospheres were derived from animals ubiquitously expressing GFP driven off an ubiquitous promoter [21]. When neurospheres were grown in co-culture with MSCs the neural cells rapidly migrated from the spheres onto the surrounding substrate.…”

Section: Discussionmentioning

confidence: 99%

Human Mesenchymal Stem Cells Signals Regulate Neural Stem Cell Fate

et al. 2006

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Neural stem cells (NSCs) differentiate into neurons, astrocytes and oligodendrocytes depending on their location within the central nervous system (CNS). The cellular and molecular cues mediating end-stage cell fate choices are not completely understood. The retention of multipotent NSCs in the adult CNS raises the possibility that selective recruitment of their progeny to specific lineages may facilitate repair in a spectrum of neuropathological conditions. Previous studies suggest that adult human bone marrow derived mesenchymal stem cells (hMSCs) improve functional outcome after a wide range of CNS insults, probably through their trophic influence. In the context of such trophic activity, here we demonstrate that hMSCs in culture provide humoral signals that selectively promote the genesis of neurons and oligodendrocytes from NSCs. Cell-cell contacts were less effective and the proportion of hMSCs that could be induced to express neural characteristics was very small. We propose that the selective promotion of neuronal and oligodendroglial fates in neural stem cell progeny is responsible for the ability of MSCs to enhance recovery after a wide range of CNS injuries.

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Analysis of Green Fluorescent Protein Expression in Transgenic Rats for Tracking Transplanted Neural Stem/Progenitor Cells

Cited by 45 publications

References 34 publications

New lines of GFP transgenic rats relevant for regenerative medicine and gene therapy

New lines of GFP transgenic rats relevant for regenerative medicine and gene therapy

Sensitive detection of GFP utilizing tyramide signal amplification to overcome gene silencing

Human Mesenchymal Stem Cells Signals Regulate Neural Stem Cell Fate

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