C57BL6 mouse substrains demonstrate differences in susceptibility to the demyelinating effects of Cuprizone toxin

Advances in our understanding of cellular functions and phenotypes in the brain rely on technically robust experimental in vivo models with face validity towards human disease. The cuprizone toxin-induced demyelination model is widely used to investigate pathophysiological mechanisms of demyelinating and remyelinating phases of multiple sclerosis. The C57BL6 mouse is a common inbred strain used as the genetic background for genetically engineered and congenic mice. Substrains of C57BL6 mice sourced from distinct vendors are often treated as equivalent in research studies. Here, we demonstrated that an alternative dosing approach via oral gavage with a well-tolerated, lower dose of cuprizone resulted in significant differences in C57BL/6NTac (Taconic) over C57BL/6J (Jax) mice. With consistent dosing of cuprizone for 5 weeks, body weights were significantly affected in C57BL/6NTac versus C57BL/6J mice. DT-MRI showed significant demyelination in white matter regions in the C57BL/6NTac mice. Concomitantly, histology analysis illustrated increased microgliosis and proliferation in C57BL/6NTac compared with C57BL/6J mice. These observations suggest that the C57BL/6NTac substrain of C57BL6 mice is more vulnerable to cuprizone challenge. Genetic factors along with breeder source appear to influence susceptibility to cuprizone toxin. Thus, the awareness of the limitations of in vivo models in addition to informed decision making on the appropriate background substrain can greatly improve sensitivity and reproducibility of results and use for evaluating investigational therapeutics.

Evaluating spiny mice (Acomys) as a model for cardiac research

Complex tissue regeneration is extremely rare among adult mammals. An exception, however, is the superior tissue healing of multiple organs in spiny mice (Acomys). While Acomys species exhibit the remarkable ability to heal complex tissue with minimal scarring, little is known about their cardiac structure and function. In this study, we characterized cardiac structure, anatomy, and in vivo function, as well as cardiomyocyte characteristics in Acomys compared to the most commonly used cardiac mouse model, the C57BL6 mouse strain (Mus). Our results demonstrate comparable cardiac anatomy, structure and function between the two rodent species, but reveal significant differences in their cardiomyocyte characteristics. These findings establish Acomys as a new mammalian model for cardiac research.

Abstract 40: Renin is Produced By a General Lysosomal Degradation Mechanism

Renin is secreted almost exclusively by the juxtaglomerular (JG) cells of the kidney where it is first made as an inactive precursor called prorenin. Conversion of prorenin to active renin requires the proteolytic removal of an N-terminal prosegment by a second protease whose identity is still debated. Active renin is then stored in dense vesicles and secreted in response to stimuli. Using confocal microscopy we found that C57BL6 mouse kidney JG cells are highly enriched in Lamp-1, a biomarker for lysosomes, and that renin and Lamp-1 co-localize in renin storage vesicles. These data suggest that renin is stored in secretory lysosomes. N-terminal sequencing of C57BL6 mouse Ren-1 renin purified from kidney revealed an N-terminus beginning with SPVVLT¼. This is the same amino terminus as that reported for rat renin and mouse As4.1 cells and different from that reported for human renin. This result raises the possibility that rodents and humans use different prorenin processing enzymes (PPE). Treatment of mice with captopril for 7 days increases plasma active renin by 19-fold (control 149 +/- 22 vs. treated 2859 +/- 672 ng AI/ml/hr, P< 0.0001) and kidney renin messenger RNA by 4.81-fold (P< 0.0001). Nevertheless, Illumina expression array analysis of C57BL6 mouse kidney before and after captopril treatment did not reveal candidate PPEs whose expression paralleled that of renin. This result suggests that the PPE is not limited to JG cells. To test the possibility that general lysosomal hydrolases are responsible for renin production, we used a Lamp-1 C-terminal sequence to force the sorting of mouse Ren-1 prorenin into lysosomes of transfected human embryonic kidney (HEK) cells. Transfection resulted in the intracellular retention of renin of the appropriate molecular weight and which lacked the engineered Lamp-1 C-terminal tail, suggesting that the proteolytic processing of prorenin is not carried out by a protease that is restricted to JG cells. Altogether, our results are consistent with mature renin being produced by lysosomal degradation of the prosegment and the selective resistance of mature renin to hydrolysis. The different N-termini of rodent and human renins could be explained by differential susceptibility of their prosegments to lysosomal hydrolysis.

Biomechanical and Microstructural Analysis of Wildtype (C57BL6) Mouse Aorta

It is generally accepted that the formation of an aneurysm in the infrarenal aorta is a complex and multi-factorial disease, however little is known about how biomechanical factors may play a role in the progression of aneurysmal disease. Although it is known that human aneurysmal tissue is remodeled in the disease process [1] and that such reorganization leads to altered function [2], the underlying mechanisms by which such changes remains an important unanswered question in the literature. The purpose of this study is to develop a means for determining the biomechanical alterations that occur within the aorta to better understand aneurysmal disease progression.