Novel phenotypes identified by plasma biochemical screening in the mouse

Abstract

We used ENU mutagenesis in the mouse for the rapid generation of novel mutant phenotypes for both gene function studies and use as new animal models of human disease (Nolan et al. 2000b). One focus of the program was the development of a blood biochemistry screen. At 8-12 weeks of age, approximately 300 ml of blood was collected from F1 offspring of ENU mutagenized male mice. This yielded approximately 125 ml of plasma, used to perform a profile of 17 standard biochemical tests on an Olympus analyzer. Cohorts of F1 mice were also aged and then retested to detect late onset phenotypes. In total, 1,961 F1s were screened. Outliers were identified by running means and standard deviations. Of 70 mice showing consistent abnormalities in plasma biochemistry, 29 were entered into inheritance testing. Of these, 9 phenotypes were confirmed as inherited, 10 found not to be inherited, and 10 are still being tested. Inherited mutant phenotypes include abnormal lipid profiles (low total and HDL cholesterol, high triglycerides); abnormalities in bone and liver metabolism (low ALP, high ALP, high ALT, and AST); abnormal plasma electrolyte levels (high sodium and chloride); as well as phenotypes of interest for the study of diabetes (high glucose). The gene loci bearing the mutations are currently being mapped and further characterized. Our results have validated our biochemical screen, which is applicable to other mutagenesis projects, and we have produced a new set of mutants with defined metabolic phenotypes.

Standard reporting requirements for biological samples in metabolomics experiments: mammalian/in vivo experiments

“…From the toxicology field standard operating protocols (SOPs) were provided from Pfizer, GlaxoSmithKline and Bayer as well as two recent peer reviewed publications from the toxicology field (Mortishire-Smith et al 2004;Poon et al 2005). For functional genomic studies to specifically address the influence of the strain background two papers were chosen that demonstrate the profound influence the genetic background has on the metabolic function and phenotype of a genetically modified organism (Hough et al 2002;Sam et al 2001). Finally a combination of clinical trials and studies of human metabolism where samples have been taken under less strict control of external variables were also considered (Brindle et al 2002;Poston et al 2006;Wolff et al 2004;Siu et al 2006;Townsley et al 2006;Khor et al 2006;Teahan et al 2006).…”

A New Mouse Model of Type 2 Diabetes, Produced by N-Ethyl-Nitrosourea Mutagenesis, Is the Result of a Missense Mutation in the Glucokinase Gene

“…ENU mutagenesis. Procedures used for ENU mutagenesis have been described previously (16,19). Identification of mice with blood glucose phenotypes.…”

“…Identification of mice with blood glucose phenotypes. Plasma biochemistry screening procedures that were used to identify mice between 8 and 12 weeks of age with glucose phenotypes in a cohort of ENU mutagenized mice have been described previously (19). Intraperitoneal glucose tolerance test.…”

“…These point mutations can create a variety of types of allele, including loss of function, hypomorphs, hypermorphs, and gain of function. This approach allows the discovery of gene function and dissection of protein functional domains in an unbiased manner (15).Here we report identification of a mouse model of the MODY form of diabetes from a dominant phenotypedriven screen of ENU mutant mice in the Harwell/SB mutagenesis project (16,19) and genetic mapping and positional cloning of the underlying genetic defect. This is the first ENU-derived mouse model of diabetes and specifically of MODY2.…”

“…Here we report identification of a mouse model of the MODY form of diabetes from a dominant phenotypedriven screen of ENU mutant mice in the Harwell/SB mutagenesis project (16,19) and genetic mapping and positional cloning of the underlying genetic defect. This is the first ENU-derived mouse model of diabetes and specifically of MODY2.…”