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A Phenotyping Regimen for Genetically Modified Mice Used to Study Genes Implicated in Human Diseases of Aging
Published on: July 14, 2016
Characterization of New Transgenic Mouse Models for Two Charcot-Marie-Tooth-Causing HspB1 Mutations using the Rosa26
Delphine Bouhy1, Thomas Geuens1, Vicky De Winter1
1Peripheral Neuropathy Group, VIB Department of Molecular Genetics and Institute Born Bunge, University of Antwerp, Antwerpen, Belgium.
Background:
Charcot-Marie-Tooth (CMT) and associated neuropathies, the most common inherited diseases of the peripheral nervous system, remain so far incurable. Three existing murine models of Charcot-Marie-Tooth type 2F (CMT2F) and/or distal hereditary motor neuropathy type IIb (dHMNIIb), caused by mutations in the small heat shock protein B1 gene (HSPB1/HSP27), partially recapitulate the hallmarks of peripheral neuropathy. Because these models overexpress the HSPB1 mutant proteins they differ from the patients' situation.
Objective:
To overcome the possible bias induced by overexpression, we generated and characterized a transgenic model in which the wild type or mutant HSPB1 protein was expressed at a moderate, more physiologically relevant level.
Methods:
We generated a new transgenic mouse model in which a human wild type (hHSPB1WT) or mutant (hHSPB1R127W; hHSPB1P182L) HSPB1 transgene was integrated in the mouse ROSA26 locus. The motor and sensory functions of the mice was assessed at 3, 6, 9, 12 and 18 month.
Results:
However, the mice expressing the mutant hHSPB1 do not develop motor or sensory deficits and do not show any sign of axonal degeneration, even at late age. Quantitative PCR analyses reveal contrasting tissue-specific expression pattern for the endogenous mouse and exogenous human HSPB1 and show that the ratio of human HSPB1 to the endogenous mouse HspB1 is lower in the sciatic nerve and spinal cord compared to the brain.
Conclusion:
These results suggest that expressing the transgene at a physiological level using the ROSA26 locus may not be sufficient to model inherited peripheral neuropathies caused by mutation in HSPB1.
Insights
New transgenic mice models for Charcot-Marie-Tooth (CMT) were created by expressing HSPB1 at physiological levels. However, these models did not develop neuropathy, suggesting this approach may not fully replicate the disease.
Area of Science:
- Neuroscience
- Genetics
- Molecular Biology
Background:
- Charcot-Marie-Tooth (CMT) and related neuropathies are common inherited peripheral nervous system disorders with no cure.
- Existing mouse models for CMT2F/dHMNIIb, caused by HSPB1 mutations, overexpress mutant proteins, differing from patient conditions.
- This overexpression may introduce bias, limiting the models' accuracy in representing human disease.
Purpose of the Study:
- To develop a more accurate transgenic mouse model for HSPB1-related neuropathies.
- To express wild-type or mutant human HSPB1 (hHSPB1) at physiologically relevant levels.
- To overcome limitations of existing models that overexpress mutant HSPB1.
Main Methods:
- Generated transgenic mice with hHSPB1WT or hHSPB1R127W/P182L transgenes integrated into the ROSA26 locus.
- Assessed motor and sensory functions in mice at multiple time points (3-18 months).
- Utilized quantitative PCR to analyze tissue-specific expression patterns of endogenous and exogenous HSPB1.
Main Results:
- Mice expressing mutant hHSPB1 did not exhibit motor/sensory deficits or axonal degeneration.
- Expression levels varied tissue-specifically, with lower human HSPB1 to mouse HspB1 ratios in the sciatic nerve and spinal cord compared to the brain.
- The ROSA26 locus integration resulted in moderate, physiological expression levels.
Conclusions:
- Expressing HSPB1 transgenes at physiological levels via the ROSA26 locus may be insufficient to model HSPB1-related inherited peripheral neuropathies.
- This suggests that higher expression or alternative modeling strategies might be needed.
- Further research is required to develop accurate preclinical models for these debilitating neurological disorders.
