| 1. |
- Englund, Ulrica, et al.
(författare)
-
The use of a recombinant lentiviral vector for ex vivo gene transfer into the rat CNS
- 2000
-
Ingår i: NeuroReport. - : Lippincott Williams & Wilkins. - 0959-4965 .- 1473-558X. ; 11:18, s. 3973-3977
-
Tidskriftsartikel (refereegranskat)abstract
- A major obstacle in ex vivo gene transfer has been the loss of transgene expression soon after implantation of the grafted transduced cells. Recently, a lentiviral vector system has been developed which has proven to express high levels of transgenes in vivo after direct injection into the tissue. In this study, we have investigated the use of such a vector for ex vivo gene transfer to the brain. A number of neural cell types were found to be permissive to transduction by the lentiviral vector in vitro and a majority of them expressed the transgene after transplantation to the rat brain. Transgene expression was detected up to 8 weeks post-grafting. These findings suggest that recombinant lentiviral vectors may be used for further development of ex vivo gene therapy protocols to the CNS.
|
|
| 2. |
- Ericson, Cecilia, et al.
(författare)
-
Ex vivo and in vitro studies of transgene expression in rat astrocytes transduced with lentiviral vectors.
- 2002
-
Ingår i: Experimental Neurology. - : Elsevier BV. - 0014-4886. ; 173:1, s. 22-30
-
Tidskriftsartikel (refereegranskat)abstract
- Implantation of cells genetically modified to express therapeutic genes into the brain has been proposed as a potential treatment for neurodegenerative diseases. In the current study embryonic rat-derived astrocytes were cultured and transduced with a lentiviral vector expressing the reporter gene green fluorescent protein (GFP) and subsequently grafted into the adult rat brain. The proportion of GFP expressing cells was stable, albeit small (1%), at all survival times, up to 6 weeks, the longest time point studied. In parallel in vitro studies, the astrocytes were lentivirally transduced to express either one of the two isoforms of glutamate decarboxylase (GAD(65) or GAD(67)) or glial cell line-derived neurotrophic factor (GDNF). When transducing 293T cells with the two GAD vectors, released GABA could be measured using high-performance liquid chromatography. Further studies of rat astrocytes transduced with the same vectors resulted in a level of GAD activity about 10 times higher than the activity of an intact rat striatum. One hundred thousand astrocytes transduced with LV-GDNF released approximately 27 ng of GDNF per hour. Thus, taken together, our observations provide support for the use of rat astrocytes in ex vivo gene transfer of these proteins in animal models of CNS disorders, e.g., Parkinson's disease or epilepsy.
|
|
| 3. |
- Ericson, Cecilia, et al.
(författare)
-
Ex vivo gene delivery of GDNF using primary astrocytes transduced with a lentiviral vector provides neuroprotection in a rat model of Parkinson's disease.
- 2005
-
Ingår i: European Journal of Neuroscience. - : Wiley. - 1460-9568 .- 0953-816X. ; 22:11, s. 2755-2764
-
Tidskriftsartikel (refereegranskat)abstract
- Astrocytes are, as normal constituents of the brain, promising vehicles for ex vivo gene delivery to the central nervous system. In the present study, we have used a lentiviral vector encoding glial cell line-derived neurotrophic factor (GDNF) to transduce rat-derived primary astrocytes, in order to evaluate their potential for long-term transgene expression in vivo and neuroprotection in a rat model of Parkinson's disease. Following transplantation of GDNF-transduced astrocytes to the intact striatum, the level of released GDNF was 2.93 +/- 0.28 ng/mg tissue at 1 week post-grafting, reduced to 0.42 +/- 0.12 ng/mg tissue at 4 weeks, and thereafter was maintained at this level throughout the experiment (12 weeks; 0.53 +/- 0.068 ng/mg tissue). Similarly, grafting to the substantia nigra (SN) resulted in a significant overexpression of GDNF ( approximately 0.20 ng/mg tissue) at 1 week. Intact animals receiving transplants of GDNF-transduced astrocytes displayed an increased contralateral turning (5.39 +/- 1.19 turns/min) in the amphetamine-induced rotation test, which significantly correlated with the GDNF tissue levels measured in the striatum, indicating a stimulatory effect of GDNF on the dopaminergic function. Transplantation of GDNF-transduced astrocytes to the SN 1 week prior to an intrastriatal 6-hydroxydopamine lesion provided a significant protection of nigral tyrosine hydroxylase-positive cells. By contrast, when the cells were transplanted to the striatum, the level of released GDNF was not sufficient to rescue the striatal fibers and, hence, to protect the nigral dopaminergic neurons. Overall, our results suggest that genetically modified astrocytes expressing GDNF can provide neuroprotection in a rat model of Parkinson's disease following transplantation to the SN.
|
|
| 4. |
|
|
| 5. |
- Georgievska, Biljana, et al.
(författare)
-
Regulated delivery of glial cell line-derived neurotrophic factor into rat striatum, using a tetracycline-dependent lentiviral vector.
- 2004
-
Ingår i: Human Gene Therapy. - : Mary Ann Liebert Inc. - 1043-0342 .- 1557-7422. ; 15:10, s. 934-944
-
Tidskriftsartikel (refereegranskat)abstract
- In this study, a tetracycline-regulated lentiviral vector system, based on the tetracycline-dependent transactivator rtTA2S-M2, was developed for controlled expression of glial cell line-derived neurotrophic factor (GDNF) in the rat brain. Expression of the marker gene green fluorescent protein (GFP) and GDNF was tightly regulated in a dose-dependent manner in neural cell lines in vitro. Injection of high-titer lentiviral vectors into the rat striatum resulted in a 7-fold induction of GDNF tissue levels (1060 pg/mg tissue), when doxycycline (a tetracycline analog) was added to the drinking water. However, low levels of GDNF (150 pg/mg tissue) were also detected in animals that did not receive doxycycline, indicating a significant background leakage from the vector system in vivo. The level of basal expression was markedly reduced when a 10-fold lower dose of the tetracycline-regulated GDNF vector was injected into the striatum (3–11 pg/mg tissue), and doxycycline- induced GDNF tissue levels obtained in these animals were about 190 pg/mg tissue. Doxycycline-induced expression of GDNF resulted in a significant downregulation of the tyrosine hydroxylase (TH) protein in the intact striatum. Removal of doxycycline from the drinking water rapidly (within 3 days) turned off transgenic GDNF mRNA expression and GDNF protein levels in the tissue were completely reduced by 2 weeks, demonstrating the dynamics of the system in vivo. Accordingly, TH protein expression returned to normal by 2–8 weeks after removal of doxycycline, indicating that GDNF-induced downregulation of TH is a reversible event.
|
|
| 6. |
- Jakobsson, Johan, et al.
(författare)
-
Lentiviral vectors.
- 2003
-
Ingår i: International Review of Neurobiology. - 0074-7742. ; 55, s. 111-122
-
Forskningsöversikt (refereegranskat)
|
|
| 7. |
- Jakobsson, Johan, et al.
(författare)
-
Lesion-dependent regulation of transgene expression in the rat brain using a human glial fibrillary acidic protein-lentiviral vector.
- 2004
-
Ingår i: European Journal of Neuroscience. - 1460-9568. ; 19:3, s. 761-765
-
Tidskriftsartikel (refereegranskat)abstract
- The ability to regulate transgene expression will be crucial for development of gene therapy to the brain. The most commonly used systems are based on a transactivator in combination with a drug, e.g. the tetracycline-regulated system. Here we describe a different method of transgene regulation by the use of the human glial fibrillary acidic protein (GFAP) promoter. We constructed a lentiviral vector that directs transgene expression to astrocytes. Using toxin-induced lesions we investigated to what extent transgene expression could be regulated in accordance with the activation of the endogenous GFAP gene. In animals receiving excitotoxic lesions of the striatum we detected an eightfold increase of green fluorescent protein (GFP)-expressing cells. The vast majority of these cells did not divide, suggesting that the transgene was indeed regulated in a similar fashion as the endogenous GFAP gene. This finding will lead to the development of lentiviral vectors with autoregulatory capacities that may be very useful for gene therapy to the brain.
|
|
| 8. |
- Jakobsson, Johan, et al.
(författare)
-
Targeted transgene expression in rat brain using lentiviral vectors.
- 2003
-
Ingår i: Journal of Neuroscience Research. - : Wiley. - 1097-4547 .- 0360-4012. ; 73:6, s. 876-885
-
Tidskriftsartikel (refereegranskat)abstract
- Direct gene transfer to the adult brain is dependent on vectors that transduce non-dividing cells, such as lentiviral vectors. Another aspect of the development of gene therapy to the brain is the need for cell-specific transgene expression. Expression from vesicular stomatitis virus G-protein (VSV-G) pseudotyped lentiviral vectors has been reported to be mainly neuron specific in the brain. We constructed cell-specific lentiviral vectors using the neuron-specific enolase (rNSE) or the glial fibrillary acidic protein (hGFAP) promoters and compared them to the ubiquitous human cytomegalovirus promoter (hCMV), a hybrid CMV/-actin promoter (CAG) and the promoter for human elongation factor 1 (EF1). Our results showed that the hGFAP promoter was expressed only in glial cells, whereas rNSE was purely neuron specific, showing that VSV-G is pantropic in the rat striatum. We conclude that the VSV-G allows transduction of both glial and neuronal cells and the promoter dictates in what cell type the transgene will be expressed. The expression of transgenes exclusively in astrocytes would allow for local delivery of secreted transgene products, such as glial cell line-derived neurotrophic factor (GDNF), circumventing the anterograde transport that may induce unwanted side effects.
|
|
| 9. |
|
|
| 10. |
- Berglind, Johanna, et al.
(författare)
-
Allt åt alla för alltid : Förslag till ett nationellt ramverk för gallring och bevarande
- 2020
-
Rapport (populärvet., debatt m.m.)abstract
- Kungliga biblioteket (KB) tillsatte sommaren 2019 en arbetsgrupp med uppdrag attpåbörja arbetet med nationella riktlinjer för gallring. Via tidigare nätverksmöten ochkonferenser samt i rapporter hade det framkommit en önskan om ett nationelltsamarbete kring gallring och bevarande.Forskningsbiblioteken (lärosätesbibliotek och specialbibliotek) gallrar i dagslägetmycket tryckt material, men arbetet sker utan samordning. Viktigt material riskerardärför att gå förlorat. Tidigare svenska samarbeten kring bevarande och gallring visar pågod vilja och goda intentioner. Dock har det funnits brister i långsiktighet ochorganisation, oklar finansiering på lång sikt och svag samordning och uppföljning. Flerapågående och framgångsrika internationella initiativ visar på behovet av samsyn kringgrundläggande mål med arbetet och en långsiktigt hållbar lösning, såväl ideologiskt somfinansiellt och organisatoriskt.Arbetsgruppen har genom omvärldsbevakning och avstämning med fältet tydliggjortkärnan i vad ett långsiktigt samarbete skulle innebära. Detta har konkretiserats i ettförslag till en vision och policy i form av ett ramverk. Visionen sammanfattas i att”forskningsbiblioteken samverkar långsiktigt och tar ett gemensamt ansvar för att spara,bevara och tillgängliggöra material för dagens och framtidens behov för alla sinamålgrupper”.Visionen konkretiseras i sex policy-punkter om samverkan, bevarande ochtillgängliggörande. Samverkan bör ske genom en formaliserad organisationsstruktur ochge förutsättningar för att se samlingarna som en gemensam resurs, så att varje bibliotekinte behöver spara allt. Minst ett exemplar av det svenska trycket ska bevaras som"samhällets minne" och ett tillräckligt antal exemplar måste sparas för att ävensäkerställa nyttjande för framtiden. Material som sparas måste tillgängliggöras effektivt(genom till exempel digitalisering och fjärrlån) och god sökbarhet upprätthålls genomkvalitativ metadata.Ramverket har diskuterats med fältet på flertalet workshoppar, vid konferensenSamlingshantering i teori och praktik1 samt genom en enkät till berörda bibliotek.Dialogen visar på ett mycket stort intresse hos biblioteken för att samarbeta kringgallring och bevarande av lågfrekvent använt material och på så sätt säkra tillgången tillforskningsmaterial. Ramverket är ett sätt att samordna resurser för att bygga relevantatjänster och säkra tillgång till samlingar för framtiden. Det krävs ett tätt samarbete ochgemensamt ansvarstagande mellan KB och forskningsbiblioteken för att ge tyngd ochförutsättningar för ett långsiktigt och hållbart arbete. Behoven och möjligheterna för biblioteken att delta i ett samarbete ser dock olika ut. Därför är det viktigt att de kandelta efter sina specifika förutsättningar.Arbetsgruppens rekommendation är att KB i det första skedet bör leda arbetet med attfastställa ramverket och dess vision med intresserade forskningsbibliotek, genomexempelvis en avsiktsförklaring. Man behöver också skapa förutsättningar förimplementering av ramverket genom att till exempel definiera roller och samordna medandra pågående utredningar. I nästa fas tas en organisation för samarbetet framgemensamt av deltagande parter. Därefter kan det praktiska arbetet starta, till exempel iform av pilotprojekt som kan utgå från bibliotekens pågående eller planeradegallringsarbeten. Detta för att i mindre skala pröva ramverket, dess finansiering ochorganisation samt framtagna riktlinjer, innan det skalas upp. Slutligen kan projektetövergå till förvaltning, där organisation, roller, finansiering och riktlinjer fastställs.
|
|
