Synchrotron radiation for preclinical research:
investigation of tissue chemical change in mouse model
of Pompe disease (glycogenosis type II) and assessment
of gene therapy efficacy by using Infrared Micro
spectroscopy with the synchrotron light.

Laurence Dubreil, Juliette Hordeaux, Johan Deniaud, Lydie Lagalice, Karim Bey, Christophe Sandt, Frédéric Jamme, Marie-Anne Colle

► To cite this version:

Laurence Dubreil, Juliette Hordeaux, Johan Deniaud, Lydie Lagalice, Karim Bey, et al. Synchrotron radiation for preclinical research: investigation of tissue chemical change in mouse model of Pompe disease (glycogenosis type II) and assessment of gene therapy efficacy by using Infrared Micro spectroscopy with the synchrotron light. Sfμ 2017, Société Française des Microscopies (SFμ). FRA., Jul 2017, Bordeaux, France. hal-01567212

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Submitted on 5 Jun 2020

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**Synchrotron radiation for preclinical research: investigation of tissue chemical change in mouse model of Pompe disease (glycogenosis type II) and assessment of gene therapy efficacy by using Infrared Micro spectroscopy with the synchrotron light.**

Laurence Dubreil<fnref n="1" />,<fnref n="2" />, Juliette Hordeaux<fnref n="1" />,<fnref n="2" />,<fnref n="3" />, Johan Deniaud<fnref n="1" />,<fnref n="2" />, Lydie Lagalice<fnref n="1" />,<fnref n="2" />, Karim Bey<fnref n="1" />,<fnref n="2" />, Christophe Sandt<fnref n="4" />, Frédéric Jamme<fnref n="4" /> and Marie-Anne Colle<fnref n="1" />,<fnref n="2" />

Pompe disease (glycogenosis type II) is an autosomal recessive disorder caused by the deficiency of the lysosomal enzyme acid alpha-glucosidase (GAA). The disease is characterized by lysosomal glycogen storage in heart and muscles, and manifests as a fatal cardiomyopathy in infantile form. Cardiac correction by enzyme replacement therapy (ERT) has recently prolonged the lifespan of these patients but ERT is not efficient to correct central nervous system (CNS). A gene therapy strategy using AAV vectors delivered to cerebrospinal fluid has been set up to restore GAA activity into the CNS. We demonstrate the use of Infrared Micro spectroscopy with synchrotron light as an innovative tool to map glycogen at the subcellular level in motor neurons (MNs) from spinal cord and cardiac fibers. Principal Component Analysis of infrared spectral data from MNs and cardiac fibers show that both treated and wild-type animals are merged in the same cluster whereas infrared spectra obtained from untreated Pompe mice are characterized by increase of the bands assigned to the carbohydrates of glycogen. This new analytical approach that allows an highly sensitive and resolutive direct probing of tissue glycogen is required to explore early biochemical change at a subcellular level and therefore to assess therapeutic efficiency for Pompe disease.

**Keywords** : Pompe disease, glycogen, Infrared Micro spectroscopy

**Acknowledgments** : We thank the vector core of the Atlantic Gene Therapies Institute (AGT) in Nantes for the preparation of the rAAV vectors, Véronique Blouin and Philippe Moullier (INSERM UMR1089) for vector production and the technical staff of Oniris rodent facility for animal care. We acknowledge assistance from SOLEIL SMIS beamline staff for his help. This work was supported by - NeurATRIS: A Translational Research Infrastructure for Biotherapies in Neurosciences. Thank to Sfµ for its financial support to participate at this congress.

<figure><img src="image_2.png" /><figcaption>**Figure 1** : a- Representative sections of cross-sectioned cervical spinal cord, paraffin embedding, PAS-luxol fast blue stain. The glycogen storage appears purple on a blue background, motor neuron of spinal cord are located in ventral horn (vh). b- Chemical imaging performed after the infrared chemical mapping showing the relative concentration of glycogen infrared spectra absorption acquired from cross-section cervical spinal cord of the dewaxed formalin fixed paraffin.</figcaption></figure>

<footnote n="1">1 INRA UMR703 PAnTher, F-44307 Nantes, France</footnote>

<footnote n="2">2 LUNAM Université, Oniris, École nationale vétérinaire, agro-alimentaire et de l'alimentation Nantes-Atlantique, Nantes, F-44307, France</footnote>

<footnote n="3">3 Université de Nantes, Nantes, France</footnote>

<footnote n="4">4 SOLEIL French national synchrotron facility, Gif-sur-Yvette, France</footnote>