Wolf Lab - Research

Rare Monogenic Diseases

Pig Models for Dystrophinopathies

Duchenne muscular dystrophy (DMD) is a severe X-linked neuromuscular disorder affecting approximately 1 in 5,000–6,000 male births. Mutations disrupting the reading frame of the DMD gene result in the absence of functional dystrophin, causing progressive muscle degeneration, loss of mobility, and ultimately respiratory and cardiac failure. Large animal models are essential for understanding disease mechanisms and evaluating emerging therapies (1; Fig. 1).

Fig. 1. Multiple Research Directions Using Tailored Pig Models for Dystrophinopathies (created with BioRender.com).

We developed a porcine DMD model lacking exon 52 (DMDΔ52), which closely reproduces the molecular, clinical, and pathological features of human DMD. Its accelerated disease progression facilitates preclinical therapeutic studies (2). The model has enabled multi-omics investigations of disease mechanisms and validation of multispectral optoacoustic tomography (MSOT) for non-invasive monitoring of muscle fibrosis (3).

Using CRISPR/Cas9-mediated deletion of exon 51, we demonstrated restoration of the DMD reading frame and expression of an internally truncated, partially functional dystrophin protein.

Systemic gene editing improved skeletal and cardiac muscle pathology and function in DMDΔ52 pigs (4). To assess the full therapeutic potential of this approach, we generated DMDΔ51-52 pigs, which exhibit a milder, Becker muscular dystrophy-like phenotype with substantially improved muscle pathology and normalized cardiac function (5). However, quantitative muscle force measurements revealed incomplete functional recovery, highlighting their importance for evaluating therapeutic efficacy (6).

Recent analyses of microRNA–mRNA networks further demonstrated partial restoration of disrupted structural, metabolic, and signaling pathways in DMDΔ51-52 pigs and identified potential biomarkers for disease stratification and therapeutic monitoring (7).

Current research focuses on inflammatory mechanisms and genetic modifiers of DMD progression, including the role of the NLRP3 inflammasome and NOTCH3. We are also evaluating a broadly applicable gene-editing strategy targeting DMD exons 45–55, with the potential to benefit a substantial proportion of DMD patients.

Pig Model for Laron Syndrome

Laron syndrome (LS) is a rare genetic disorder caused by loss-of-function mutations in the growth hormone receptor (GHR) gene, resulting in growth hormone (GH) insensitivity and severely reduced insulin-like growth factor 1 (IGF1) levels. To model LS, GHR-knockout (KO) pigs were generated using CRISPR/Cas9 gene editing (8). These pigs closely reproduce the hormonal, growth, and metabolic characteristics of human LS.

Growth and body composition: GHR-KO pigs exhibit markedly reduced IGF1 and IGFBP3, elevated GH and IGFBP2 levels, and altered pituitary GH production and secretion (8, 9). Although birth weight is normal, growth retardation becomes apparent by five weeks, resulting in approximately 60% lower body weight at six months. Most organs are smaller, particularly the liver, kidneys, and heart, while relative brain weight increases. These characteristics also make GHR-KO pigs relevant for xenotransplantation research (10). Increased body fat, adipocyte hypertrophy, and depot-specific adipose tissue changes closely resemble those observed in human LS (11).

Glucose metabolism and pancreatic function: GHR-KO pigs develop transient juvenile hypoglycemia despite markedly reduced pancreatic β-cell volume (83% in young and 73% in adult pigs) and impaired insulin secretion (12, 13). Nevertheless, glucose tolerance remains preserved due to increased insulin sensitivity, as confirmed by hyperinsulinemic-euglycemic clamp studies (14; Fig. 2). These findings highlight the importance of GH signaling for β-cell development and glucose homeostasis.

Fig. 2. Mechanisms leading to juvenile hypoglycemia in GHR deficiency and its normalization during puberty (from Ref. 14; created with BioRender.com).

Liver signaling and metabolism: Loss of GHR reduces hepatic STAT5 activation while increasing phosphorylation of IRS1, PI3K, JAK2, and mTOR, indicating substantial changes in intracellular signaling (8). Integrated proteomic and metabolomic analyses revealed extensive hepatic metabolic reprogramming, including enhanced amino acid degradation, urea cycle and TCA cycle activity, altered lipid metabolism, and changes in methionine and glutathione pathways. Interestingly, sex-specific differences in hepatic protein abundance observed in controls were absent in GHR-KO pigs (15).

Conclusion: GHR-KO pigs faithfully reproduce key features of human Laron syndrome, providing a valuable large-animal model to investigate GH-dependent growth, metabolic regulation, pancreatic β-cell biology, and potential therapeutic interventions.

Funding

German Research Foundation (DFG; FOR 5795: HyperMet) | German Center for Diabetes Research (DZD; 82DZD00801, 82DZD00802, 82DZD08D03, 82DZD00804, 82DZD08E1G, 82DZD08I1G) | German Center for Child and Adolescent Health (DZKJ; 01GL2406A) | Bavarian Research Foundation (AZ 802-08, PDOC-90-15) | Else Kröner-Fresenius Foundation (2015_180, 2018_T20, 2022_EKSE.41) | European Union (Innovative Health Initiative, NHPig project 101165643) | Leducq Foundation (23CVD01) | MyoGene Bio / California Institute for Regenerative Medicine (CIRM).

Selected Publications

1. Stirm M et al. Pig models for translational Duchenne muscular dystrophy research. Trends Mol Med. 2024;30:950–964. doi:10.1016/j.molmed.2024.04.013
2. Stirm M et al. A scalable, clinically severe pig model for Duchenne muscular dystrophy. Dis Model Mech. 2021;14. doi:10.1242/dmm.049285
3. Regensburger AP et al. Detection of collagens by multispectral optoacoustic tomography as an imaging biomarker for Duchenne muscular dystrophy. Nat Med. 2019;25:1905–1915. doi:10.1038/s41591-019-0669-y
4. Moretti A et al. Somatic gene editing ameliorates skeletal and cardiac muscle failure in pig and human models of Duchenne muscular dystrophy. Nat Med. 2020;26:207–214. doi:10.1038/s41591-019-0738-2
5. Stirm M et al. Systemic deletion of DMD exon 51 rescues clinically severe Duchenne muscular dystrophy in a pig model lacking DMD exon 52. PNAS. 2023;120. doi:10.1073/pnas.2301250120
6. Blasi M et al. Reduced Muscle Force in Dystrophic DMDΔ52 Pigs Is Incompletely Restored by Systemic Transcript Reframing (DMDΔ51-52). J Cachexia Sarcopenia Muscle. 2025;16. doi:10.1002/jcsm.70084
7. Reschke S et al. MicroRNA-mRNA Networks in Skeletal Muscle of Tailored Pig Models for Dystrophinopathies. J Cachexia Sarcopenia Muscle. 2026;17. doi:10.1002/jcsm.70337
8. Hinrichs A, et al. Growth hormone receptor-deficient pigs resemble the pathophysiology of human Laron syndrome and reveal altered activation of signaling cascades in the liver. Mol Metab. 2018;11:113–128. https://doi.org/10.1016/j.molmet.2018.03.006
9. Shashikadze B, et al. Structural and proteomic repercussions of growth hormone receptor deficiency on the pituitary gland: Lessons from a translational pig model. J Neuroendocrinol. 2024;36. https://doi.org/10.1111/jne.13277
10. Hinrichs A, et al. Growth hormone receptor knockout to reduce the size of donor pigs for preclinical xenotransplantation studies. Xenotransplantation. 2021;28. https://doi.org/10.1111/xen.12664
11. Young JA, et al. Growth hormone insensitivity and adipose tissue: tissue morphology and transcriptome analyses in pigs and humans. Pituitary. 2023;26:660–674. https://doi.org/10.1007/s11102-023-01355-y
12. Hinrichs A, et al. Transient juvenile hypoglycemia in growth hormone receptor deficiency – mechanistic insights from Laron syndrome and tailored animal models. Eur J Endocrinol. 2021;185–R47. https://doi.org/10.1530/EJE-21-0013
13. Laane L, et al. Decreased β-cell volume and insulin secretion but preserved glucose tolerance in a growth hormone insensitive pig model. Pituitary. 2024;27:567–576. https://doi.org/10.1007/s11102-024-01424-w
14. Hinrichs A, et al. Transient juvenile hypoglycemia in GH insensitive Laron syndrome pigs is associated with insulin hypersensitivity. Mol Metab. 2026;103:102273. https://doi.org/10.1016/j.molmet.2025.102273
15. Riedel EO, et al. Functional changes of the liver in the absence of growth hormone (GH) action – Proteomic and metabolomic insights from a GH receptor deficient pig model. Mol Metab. 2020;36:100978. https://doi.org/10.1016/j.molmet.2020.100978