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Production of KRIT1-knockout and KRIT1-knockin Mouse Embryonic Fibroblasts as Cellular Models of CCM Disease
- Source :
- Methods in Molecular Biology ISBN: 9781071606391
- Publication Year :
- 2020
- Publisher :
- Humana Press, 2020.
-
Abstract
- The development of distinct cellular and animal models has allowed the identification and characterization of molecular mechanisms underlying the pathogenesis of cerebral cavernous malformation (CCM) disease. This is a major cerebrovascular disorder of proven genetic origin, affecting 0.5% of the population. Three disease genes have been identified: CCM1/KRIT1, CCM2, and CCM3. These genes encode for proteins implicated in the regulation of major cellular structures and mechanisms, such as cell-cell and cell-matrix adhesion, actin cytoskeleton dynamics, and endothelial-to-mesenchymal transition, suggesting that they may act as pleiotropic regulators of cellular homeostasis. Indeed, accumulated evidence in cellular and animal models demonstrates that emerged pleiotropic functions of CCM proteins are mainly due to their ability to modulate redox-sensitive pathways and mechanisms involved in adaptive responses to oxidative stress and inflammation, thus contributing to the preservation of cellular homeostasis and stress defenses. In particular, we demonstrated that KRIT1 loss-of-function affects master regulators of cellular redox homeostasis and responses to oxidative stress, including major redox-sensitive transcriptional factors and antioxidant proteins, and autophagy, suggesting that altered redox signaling and oxidative stress contribute to CCM pathogenesis, and opening novel preventive and therapeutic perspectives.In this chapter, we describe materials and methods for isolation of mouse embryonic fibroblast (MEF) cells from homozygous KRIT1-knockout mouse embryos, and their transduction with a lentiviral vector encoding KRIT1 to generate cellular models of CCM disease that contributed significantly to the identification of pathogenetic mechanisms.
- Subjects :
- 0301 basic medicine
Heterozygous and homozygous KRIT1 knockout mice
Cerebral cavernous malformation (CCM)
KRIT1
Population
Cellular homeostasis
CCM genes
Cellular and animal models of CCM disease
Cerebrovascular diseases
KRIT1 knockout mouse embryonic fibroblast (MEF) cells
Biology
medicine.disease_cause
03 medical and health sciences
0302 clinical medicine
medicine
education
Transcription factor
education.field_of_study
Autophagy
Cerebrovascular disorder
Actin cytoskeleton
Embryonic stem cell
Cell biology
030104 developmental biology
030217 neurology & neurosurgery
Oxidative stress
Subjects
Details
- Language :
- English
- ISBN :
- 978-1-07-160639-1
- ISBNs :
- 9781071606391
- Database :
- OpenAIRE
- Journal :
- Methods in Molecular Biology ISBN: 9781071606391
- Accession number :
- edsair.doi.dedup.....d6261686aea5000653ef5df23e8273b8