Generation of a neurodegenerative disease mouse model using lentiviral vectors carrying an enhanced synapsin I promoter

Matsuzaki, Yasunori; Oue, Miho; Hirai, Hirokazu

doi:10.1016/j.jneumeth.2013.12.004

Cited by 26 publications

(22 citation statements)

References 44 publications

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“…21 The Syn-mCMV-FMRP vector utilized a modified hybrid promoter consisting of the human synapsin 1 gene promoter in tandem with a minimal CMV enhancer element. 24 A third vector (“null vector”) containing a CMV promoter sequence, but no transgene served as a negative control. Five treatment groups were studied: (1) WT mice injected with the null vector served as the baseline controls; (2) WT mice injected with the Syn-FMRP vector; (3) Fmr1 KO mice injected with the null vector; (4) KO mice injected with the Syn-FMRP vector; and (5) KO mice injected with the Syn-mCMV-FMRP vector.…”

Section: Resultsmentioning

confidence: 99%

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FMRP Expression Levels in Mouse Central Nervous System Neurons Determine Behavioral Phenotype

et al. 2016

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Fragile X mental retardation protein (FMRP) is absent or highly reduced in Fragile X Syndrome, a genetic disorder causing cognitive impairment and autistic behaviors. Previous proof-of-principle studies have demonstrated that restoring FMRP in the brain using viral vectors can improve pathological abnormalities in mouse models of fragile X. However, unlike small molecule drugs where the dose can readily be adjusted during treatment, viral vector–based biological therapeutic drugs present challenges in terms of achieving optimal dosing and expression levels. The objective of this study was to investigate the consequences of expressing varying levels of FMRP selectively in neurons of Fmr1 knockout and wild-type (WT) mice. A wide range of neuronal FMRP transgene levels was achieved in individual mice after intra-cerebroventricular administration of adeno-associated viral vectors coding for FMRP. In all treated knockout mice, prominent FMRP transgene expression was observed in forebrain structures, whereas lower levels were present in more caudal regions of the brain. Reduced levels of the synaptic protein PSD-95, elevated levels of the transcriptional modulator MeCP2, and abnormal motor activity, anxiety, and acoustic startle responses in Fmr1 knockout mice were fully or partially rescued after expression of FMRP at about 35–115% of WT expression, depending on the brain region examined. In the WT mouse, moderate FMRP over-expression of up to about twofold had little or no effect on PSD-95 and MeCP2 levels or on behavioral endophenotypes. In contrast, excessive over-expression in the Fmr1 knockout mouse forebrain (approximately 2.5–6-fold over WT) induced pathological motor hyperactivity and suppressed the startle response relative to WT mice. These results delineate a range of FMRP expression levels in the central nervous system that confer phenotypic improvement in fragile X mice. Collectively, these findings are pertinent to the development of long-term curative gene therapy strategies for treating Fragile X Syndrome and other neurodevelopmental disorders.

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Section: Resultsmentioning

confidence: 99%

“…The vector designated here as “Syn-FMRP” containing these domains was previously described. 21 The vector designated here as “Syn-mCMV-FMRP” also contains these domains, and a minimal cytomegalovirus (mCMV) sequence 24 was inserted at the 3′ end of the synapsin promoter, upstream of the FMRP transgene (see Fig. 1).…”

Section: Methodsmentioning

confidence: 99%

FMRP Expression Levels in Mouse Central Nervous System Neurons Determine Behavioral Phenotype

et al. 2016

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“…Cre recombinase and mtTA were delivered to the cerebellum through AAV9-SynImCMV-HA-mtTA-P2A-Cre vectors, which expressed mtTA and Cre bicistronically through a synapsin I promoter with a minimal CMV sequence (SynImCMV) (Fig 2A-1). To express mtTA and Cre in all neuronal cell types in the cerebellum, we used a neuron-specific SynImCMV promoter [36] instead of a PC-specific promoter because we had been unable to detect the fusion of hfMSCs with other neuronal cell types.…”

Section: Resultsmentioning

confidence: 99%

“…The GFP-expressing PCs co-immunostained for calbindin (Fig 3E–3G). Since we have previously found that 97% of transduced cells in the cerebellum with a neuron-specific promoter (SynIminCMV) were theoretically neurons [36], the morphological similarity of GFP-labeled cells in the molecular layer indicated that they could be basket cells (Fig 3C) or stellate cells (Fig 3D). These results strongly suggest that hfMSCs transplanted into the cerebellar cortex fuse with PCs, and presumably interneurons as well, within two weeks of transplantation.…”

Section: Resultsmentioning

confidence: 99%

“…To express transgenes specifically in PCs or various types of neurons, a PC-specific L7 promoter [35] or neuron-specific synapsin I promoter with a minimal CMV promoter sequence (SynImCMV) [36] was incorporated into the expression vector for AAV9. The mammalianized tetracycline transactivator (mtTA) cDNA [37] was kindly provided by Dr. Kenji Tanaka (Keio University).…”

Section: Methodsmentioning

confidence: 99%

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Fusion of Human Fetal Mesenchymal Stem Cells with “Degenerating” Cerebellar Neurons in Spinocerebellar Ataxia Type 1 Model Mice

et al. 2016

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Mesenchymal stem cells (MSCs) migrate to damaged tissues, where they participate in tissue repair. Human fetal MSCs (hfMSCs), compared with adult MSCs, have higher proliferation rates, a greater differentiation capacity and longer telomeres with reduced senescence. Therefore, transplantation of quality controlled hfMSCs is a promising therapeutic intervention. Previous studies have shown that intravenous or intracortical injections of MSCs result in the emergence of binucleated cerebellar Purkinje cells (PCs) containing an MSC-derived marker protein in mice, thus suggesting a fusion event. However, transdifferentiation of MSCs into PCs or transfer of a marker protein from an MSC to a PC cannot be ruled out. In this study, we unequivocally demonstrated the fusion of hfMSCs with murine PCs through a tetracycline-regulated (Tet-off) system with or without a Cre-dependent genetic inversion switch (flip-excision; FLEx). In the FLEx-Tet system, we performed intra-cerebellar injection of viral vectors expressing tetracycline transactivator (tTA) and Cre recombinase into either non-symptomatic (4-week-old) or clearly symptomatic (6–8-month-old) spinocerebellar ataxia type 1 (SCA1) mice. Then, the mice received an injection of 50,000 genetically engineered hfMSCs that expressed GFP only in the presence of Cre recombinase and tTA. We observed a significant emergence of GFP-expressing PCs and interneurons in symptomatic, but not non-symptomatic, SCA1 mice 2 weeks after the MSC injection. These results, together with the results obtained using age-matched wild-type mice, led us to conclude that hfMSCs have the potential to preferentially fuse with degenerating PCs and interneurons but not with healthy neurons.

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Methylation silencing and reactivation of exogenous genes in lentivirus-mediated transgenic mice

Wen

Cao

et al. 2021

Transgenic Res

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Generation of a neurodegenerative disease mouse model using lentiviral vectors carrying an enhanced synapsin I promoter

Cited by 26 publications

References 44 publications

FMRP Expression Levels in Mouse Central Nervous System Neurons Determine Behavioral Phenotype

FMRP Expression Levels in Mouse Central Nervous System Neurons Determine Behavioral Phenotype

Fusion of Human Fetal Mesenchymal Stem Cells with “Degenerating” Cerebellar Neurons in Spinocerebellar Ataxia Type 1 Model Mice

Methylation silencing and reactivation of exogenous genes in lentivirus-mediated transgenic mice

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