| 1. |
- Berglin-Enquist, Ida
(författare)
-
Development of Mouse Models and Gene Therapy for Gaucher Disease
- 2009
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Gaucher disease (GD) patients cannot metabolize glycosphingolipids properly due to deficiency of the enzyme glucosylceramidase (GCase). The lack of animal model for GD has hindered comprehensive investigation of disease mechanisms and also the development of curative treatment strategies such as hematopoietic stem cell (HSC) targeted gene therapy. Previous GCase deficient mice have either been lethal (due to disruption of the skin-barrier) or viable but without relevant symptoms of GD. We applied conditional strategies to create two different mouse models of GD, importantly sparing GCase activity in the skin during fetal development. Using the cre/loxP system and the Mx1-cre mouse we could develop an inducible mouse model with pathology and symptoms similar to type 1 GD patients, including infiltration of disease-characteristic Gaucher cells in target tissues, splenomegaly and anemia. We further used this model to demonstrate that HSC targeted gene therapy can cure manifest type 1 GD. The unfortunate occurrence of leukemia in otherwise successful clinical gene therapy trials has called attention to safety and by using two different transplantation-assays we provide evidence that GD has the potential to successfully respond to reduced-risk gene therapy protocols. In the second model, the function of a GBA null allele could be restored in the skin through breeding onto the K14-cre mouse, in which cre expression is restricted to the basal layer of the epidermis. This model developed early onset and rapidly progressive neurodegeneration characteristic of the severe neuronopathic form of GD. In this model we also observed considerable microglia activation. Microglia activation has been demonstrated to play an important role in the pathogenesis of other neurodegenerative conditions. Using a second mouse model in which GCase deficiency was restricted to neurons and macroglia we could demonstrate that the GCase deficient miroglia have a modulating role on disease onset and progression but are not the primary determinant of the severe neurodegeneration. Taken together the work described in this thesis provide the essential tools and proof-of-principle basis to proceed towards enhanced understanding of pathological mechanisms as well as development of novel treatment-modalities including HSC targeted gene therapy.
|
|
| 2. |
|
|
| 3. |
- Berglin-Enquist, Ida, et al.
(författare)
-
Successful Low-Risk Hematopoietic Cell Therapy in a Mouse Model of Type 1 Gaucher Disease
- 2009
-
Ingår i: Stem Cells. - : Oxford University Press (OUP). - 1549-4918 .- 1066-5099. ; 27:3, s. 744-752
-
Tidskriftsartikel (refereegranskat)abstract
- Hematopoietic stem cell-based gene therapy offers the possibility of permanent correction for genetic disorders of the hematopoietic system. However, optimization of present protocols is required before gene therapy can be safely applied as general treatment of genetic diseases. In this study we have used a mouse model of type 1 Gaucher disease (GD) to demonstrate the feasibility of a low-risk conditioning regimen instead of standard radiation, which is associated with severe adverse effects. We first wanted to establish what level of engraftment and glucosylceramidase (GCase) activity is required to correct the pathology of the type 1 GD mouse. Our results demonstrate that a median wild-type (WT) cell engraftment of 7%, corresponding to GCase activity levels above 10 nmoles/hour and mg protein, was sufficient to reverse pathology in bone marrow and spleen in the GD mouse. Moreover, we applied nonmyeloablative doses of busulfan as a pretransplant conditioning regimen and show that even WT cell engraftment in the range of 1%-10% can confer a beneficial therapeutical outcome in this disease model. Taken together, our data provide encouraging evidence for the possibility of developing safe and efficient conditioning protocols for diseases that require only a low level of normal or gene-corrected cells for a permanent and beneficial therapeutic outcome. STEM CELLS 2009; 27: 744-752
|
|
| 4. |
- Farfel-Becker, Tamar, et al.
(författare)
-
No evidence for activation of the unfolded protein response in neuronopathic models of Gaucher disease
- 2009
-
Ingår i: Human Molecular Genetics. - : Oxford University Press (OUP). - 0964-6906 .- 1460-2083. ; 18:8, s. 1482-1488
-
Tidskriftsartikel (refereegranskat)abstract
- Gaucher disease (GD), the most common lysosomal storage disorder (LSD), is caused by defects in the activity of the lysosomal enzyme, glucocerebrosidase, resulting in intracellular accumulation of glucosylceramide (GlcCer). Neuronopathic forms, which comprise only a small percent of GD patients, are characterized by neurological impairment and neuronal cell death. Little is known about the pathways leading from GlcCer accumulation to neuronal death or dysfunction but defective calcium homeostasis appears to be one of the pathways involved. Recently, endoplasmic reticulum stress together with activation of the unfolded protein response (UPR) has been suggested to play a key role in cell death in neuronopathic forms of GD, and moreover, the UPR was proposed to be a common mediator of apoptosis in LSDs (Wei et al. (2008) Hum. Mol. Genet. 17, 469-477). We now systematically examine whether the UPR is activated in neuronal forms of GD using a selection of neuronal disease models and a combination of western blotting and semi-quantitative and quantitative real-time polymerase chain reaction. We do not find any changes in either protein or mRNA levels of a number of typical UPR markers including BiP, CHOP, XBP1, Herp and GRP58, in either cultured Gaucher neurons or astrocytes, or in brain regions from mouse models, even at late symptomatic stages. We conclude that the proposition that the UPR is a common mediator for apoptosis in all neurodegenerative LSDs needs to be re-evaluated.
|
|
