Abstract
Assessing histological changes is essential for characterizing muscle disease progression and for studying the response to therapies in Duchenne muscular dystrophy (DMD), an X-linked progressive muscle-wasting disease caused by the loss of the dystrophin protein. Canine models are by far the best-characterized large animal models for DMD. In this chapter, we describe methods for muscle tissue collection and storage, hematoxylin and eosin staining for studying general muscle morphology, and special staining protocols for evaluating fibrosis, calcification, and neuronal nitric oxide synthase (nNOS) activity. We also provide immunofluorescence staining protocols that are often used to characterize the expression and localization of dystrophin and components of the dystrophin-associated glycoprotein complex. Lastly, we presented immunohistochemical staining protocols that we use to assess muscle inflammation and immune responses.
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References
Duan D, Goemans N, Takeda S et al (2021) Duchenne muscular dystrophy. Nat Rev Dis Primer 7:13
McGreevy JW, Hakim CH, McIntosh MA et al (2015) Animal models of Duchenne muscular dystrophy: from basic mechanisms to gene therapy. Dis Model Mech 8(3):195–213. https://doi.org/10.1242/dmm.018424
Wasala NB, Chen SJ, Duan D (2020) Duchenne muscular dystrophy animal models for high-throughput drug discovery and precision medicine. Expert Opin Drug Discov 15:1–14. https://doi.org/10.1080/17460441.2020.1718100
Lai Y, Thomas GD, Yue Y et al (2009) Dystrophins carrying spectrin-like repeats 16 and 17 anchor nNOS to the sarcolemma and enhance exercise performance in a mouse model of muscular dystrophy. J Clin Invest 119(3):624–635. https://doi.org/10.1172/JCI36612
Lai Y, Zhao J, Yue Y et al (2013) alpha2 and alpha3 helices of dystrophin R16 and R17 frame a microdomain in the alpha1 helix of dystrophin R17 for neuronal NOS binding. Proc Natl Acad Sci U S A 110(2):525–530. https://doi.org/10.1073/pnas.1211431109
Heckmatt JZ, Moosa A, Hutson C et al (1984) Diagnostic needle muscle biopsy. A practical and reliable alternative to open biopsy. Arch Dis Child 59(6):528–532. https://doi.org/10.1136/adc.59.6.528
Akarolo-Anthony SN, Ogundiran TO, Nkwodimmah C et al (2012) Office based muscle biopsy using Vacora vacuum assisted biopsy system. Afr J Med Med Sci 41(3):313–316
Mikell CB, Chan AK, Stein GE et al (2013) Muscle and nerve biopsies: techniques for the neurologist and neurosurgeon. Clin Neurol Neurosurg 115(8):1206–1214. https://doi.org/10.1016/j.clineuro.2013.05.008
Yue Y, Shin J-H, Duan D (2011) Whole body skeletal muscle transduction in neonatal dogs with AAV-9. Methods Mol Biol 709:313–329. https://doi.org/10.1007/978-1-61737-982-6_21
Pistolese CA, Ciarrapico AM, Della Gatta F et al (2009) Cost-effectiveness analysis of two vacuum-assisted breast biopsy systems: Mammotome and Vacora. Radiol Med 114(5):743–756. https://doi.org/10.1007/s11547-009-0404-8
Gallo A, Abraham A, Katzberg HD et al (2018) Muscle biopsy technical safety and quality using a self-contained, vacuum-assisted biopsy technique. Neuromuscul Disord 28(5):450–453. https://doi.org/10.1016/j.nmd.2018.02.006
Barthelemy F, Woods JD, Nieves-Rodriguez S et al (2020) A well-tolerated core needle muscle biopsy process suitable for children and adults. Muscle Nerve 62(6):688–698. https://doi.org/10.1002/mus.27041
Acknowledgments
The research on the canine DMD model in the Duan lab is currently supported by the National Institutes of Health (NS-90634 and AR-70517), Jesse Davidson Foundation-Defeat Duchenne Canada, Hope for Javier, Jackson Freel DMD Research Fund, Parent Project Muscular Dystrophy, Jett Foundation, Michael’s Cause, Ryan’s Quest, Solid Biosciences Inc., and the University of Missouri.
Disclosure
DD is a member of the scientific advisory board for Solid Biosciences and equity holders of Solid Biosciences. DD is a member of the scientific advisory board for Sardocor Corp.In the last 3 years, the Duan lab has received research supports unrelated to this project from Solid Biosciences and Edgewise.
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Hakim, C.H., Burke, M.J., Teixeira, J., Duan, D. (2023). Histological Assessment of Gene Therapy in the Canine DMD Model. In: Maruyama, R., Yokota, T. (eds) Muscular Dystrophy Therapeutics. Methods in Molecular Biology, vol 2587. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2772-3_16
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DOI: https://doi.org/10.1007/978-1-0716-2772-3_16
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