EYES2023 ESE Young Endocrinologists and Scientists (EYES) 2023 Oral communication 1: Adrenal Diseases (8 abstracts)
Generation and characterization of CYP21A2-I173N MICE: A humanized mutant animal model for 21-hydroxylase deficiency
Shamini Ramkumar Thirumalasetty 1 , Tina Schubert 1 , Ronald Naumann 2 , Ilka Reichardt 3 , Marie-Luise Rohm 1 , Dana Landgraf 1 , Mirko Peitzsch 4 , Mihail Sarov 3 , Nicole Reisch 5 , Angela Huebner 1 & Katrin Koehler 1
1Universitätsklinikum Carl Gustav Carus, Technische Universität Dresden, Division of Paediatric Endocrinology and Diabetes, Dresden, Germany; 2Max Planck Institute of Molecular Cell Biology and Genetics, Transgenic Core Facility, Dresden, Germany; 3Max Planck Institute of Molecular Cell Biology and Genetics, Genome Engineering Facility, Dresden, Germany; 4Universitätsklinikum Carl Gustav Carus, Technische Universität Dresden,, Institute of Clinical Chemistry and Laboratory Medicine, Dresden, Germany; 5Ludwig-Maximilians-Universität München, Medizinische Klinik und Poliklinik IV, Munich, Germany.
Background: Congenital Adrenal Hyperplasia (CAH) is a group of inherited disorders affecting adrenal steroidogenesis. The main form, 21-hydroxylase deficiency (21-OHD), results from mutations in the CYP21A2 gene. Patients experience hormone deficiencies, and excessive androgens which lead to various symptoms such as hypoglycemia, salt wasting, virilization, and early puberty. Therapy demands high glucocorticoid doses, causing significant side-effects. While new treatment approaches are emerging, there is a lack of a viable animal model for pre-clinical testing.
Objective: Development of a novel mouse model with the human CYP21A2 gene, carrying the mutation, p.I173N. A common mutation responsible for the simple virilizing form of CAH, which can also be detected in patients with the salt-wasting variant of the disease.
Methods: CRISPR-Cas9-mediated genome editing to replace the mouse Cyp21a1 gene with the human CYP21A2 gene and furthermore, to integrate the point mutation p.I173N. Heterozygote dams required dexamethasone treatment during pregnancy and until weaning. Oestrous stage was evaluated from vaginal smears at ten weeks. Mouse plasma at 20-weeks was used to study steroid hormone concentrations using LC–MS/MS. Gene-expression and histological analysis were performed using organs snap-frozen in liquid nitrogen or fixed in paraffin.
Results: At 20 weeks, homozygous mice exhibited hyperplastic adrenals and expressed the human CYP21A2 gene instead of the mouse Cyp21a1 gene. Reduced corticosterone and 11-deoxycorticosterone levels were measured in homozygotes. Progesterone levels were significantly higher (P<0.01) in homozygous mice. Homozygous mice presented with reduced concentrations of catecholamines. Male mutants exhibited normal fertility, while females were not fertile. The number of ovarian follicles were not reduced but their size was smaller and more peripheral in the persistent diestrus phase.
Conclusions: In summary, our study demonstrates that the CYP21A2 humanized mutant mice serve as a valuable model to evaluate innovative treatment approaches and will play a crucial role in amalgamating fundamental research and clinical implementation.
Volume 93
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Endocrine Abstracts
ISSN 1470-3947 (print) | ISSN 1479-6848 (online)
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