Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels
In vitro models of patient-derived muscle allow for more efficient development of genetic medicines for the muscular dystrophies, which often present mutation-specific pathologies. One popular strategy to generate patient-specific myotubes involves reprogramming dermal fibroblasts to a muscle lineag...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2022-04-01
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| Series: | Frontiers in Cell and Developmental Biology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fcell.2022.830415/full |
| _version_ | 1811297552807493632 |
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| author | Florian Barthélémy Florian Barthélémy Jeffrey W. Santoso Laura Rabichow Laura Rabichow Rongcheng Jin Isaiah Little Isaiah Little Stanley F. Nelson Stanley F. Nelson Stanley F. Nelson Megan L. McCain Megan L. McCain M. Carrie Miceli M. Carrie Miceli |
| author_facet | Florian Barthélémy Florian Barthélémy Jeffrey W. Santoso Laura Rabichow Laura Rabichow Rongcheng Jin Isaiah Little Isaiah Little Stanley F. Nelson Stanley F. Nelson Stanley F. Nelson Megan L. McCain Megan L. McCain M. Carrie Miceli M. Carrie Miceli |
| author_sort | Florian Barthélémy |
| collection | DOAJ |
| description | In vitro models of patient-derived muscle allow for more efficient development of genetic medicines for the muscular dystrophies, which often present mutation-specific pathologies. One popular strategy to generate patient-specific myotubes involves reprogramming dermal fibroblasts to a muscle lineage through MyoD induction. However, creating physiologically relevant, reproducible tissues exhibiting multinucleated, aligned myotubes with organized striations is dependent on the introduction of physicochemical cues that mimic the native muscle microenvironment. Here, we engineered patient-specific control and dystrophic muscle tissues in vitro by culturing and differentiating MyoD–directly reprogrammed fibroblasts isolated from one healthy control subject, three patients with Duchenne muscular dystrophy (DMD), and two Limb Girdle 2A/R1 (LGMD2A/R1) patients on micromolded gelatin hydrogels. Engineered DMD and LGMD2A/R1 tissues demonstrated varying levels of defects in α-actinin expression and organization relative to control, depending on the mutation. In genetically relevant DMD tissues amenable to mRNA reframing by targeting exon 44 or 45 exclusion, exposure to exon skipping antisense oligonucleotides modestly increased myotube coverage and alignment and rescued dystrophin protein expression. These findings highlight the value of engineered culture substrates in guiding the organization of reprogrammed patient fibroblasts into aligned muscle tissues, thereby extending their value as tools for exploration and dissection of the cellular and molecular basis of genetic muscle defects, rescue, and repair. |
| first_indexed | 2024-04-13T06:06:07Z |
| format | Article |
| id | doaj.art-73fc79b089c845ef877ae95d996f78e5 |
| institution | Directory Open Access Journal |
| issn | 2296-634X |
| language | English |
| last_indexed | 2024-04-13T06:06:07Z |
| publishDate | 2022-04-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Cell and Developmental Biology |
| spelling | doaj.art-73fc79b089c845ef877ae95d996f78e52022-12-22T02:59:14ZengFrontiers Media S.A.Frontiers in Cell and Developmental Biology2296-634X2022-04-011010.3389/fcell.2022.830415830415Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin HydrogelsFlorian Barthélémy0Florian Barthélémy1Jeffrey W. Santoso2Laura Rabichow3Laura Rabichow4Rongcheng Jin5Isaiah Little6Isaiah Little7Stanley F. Nelson8Stanley F. Nelson9Stanley F. Nelson10Megan L. McCain11Megan L. McCain12M. Carrie Miceli13M. Carrie Miceli14Department of Microbiology Immunology and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA, United StatesCenter for Duchenne Muscular Dystrophy, University of California, Los Angeles, Los Angeles, CA, United StatesLaboratory for Living Systems Engineering, Department of Biomedical Engineering, USC Viterbi School of Engineering, University of Southern California, Los Angeles, CA, United StatesDepartment of Microbiology Immunology and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA, United StatesCenter for Duchenne Muscular Dystrophy, University of California, Los Angeles, Los Angeles, CA, United StatesLaboratory for Living Systems Engineering, Department of Biomedical Engineering, USC Viterbi School of Engineering, University of Southern California, Los Angeles, CA, United StatesDepartment of Microbiology Immunology and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA, United StatesCenter for Duchenne Muscular Dystrophy, University of California, Los Angeles, Los Angeles, CA, United StatesCenter for Duchenne Muscular Dystrophy, University of California, Los Angeles, Los Angeles, CA, United StatesDepartment of Neurology, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA, United StatesDepartment of Pathology and Laboratory Medicine, David Geffen School of Medicine, University of California, Los Angeles, Los Angeles, CA, United StatesLaboratory for Living Systems Engineering, Department of Biomedical Engineering, USC Viterbi School of Engineering, University of Southern California, Los Angeles, CA, United StatesDepartment of Stem Cell Biology and Regenerative Medicine, Keck School of Medicine of USC, University of Southern California, Los Angeles, CA, United StatesDepartment of Microbiology Immunology and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA, United StatesCenter for Duchenne Muscular Dystrophy, University of California, Los Angeles, Los Angeles, CA, United StatesIn vitro models of patient-derived muscle allow for more efficient development of genetic medicines for the muscular dystrophies, which often present mutation-specific pathologies. One popular strategy to generate patient-specific myotubes involves reprogramming dermal fibroblasts to a muscle lineage through MyoD induction. However, creating physiologically relevant, reproducible tissues exhibiting multinucleated, aligned myotubes with organized striations is dependent on the introduction of physicochemical cues that mimic the native muscle microenvironment. Here, we engineered patient-specific control and dystrophic muscle tissues in vitro by culturing and differentiating MyoD–directly reprogrammed fibroblasts isolated from one healthy control subject, three patients with Duchenne muscular dystrophy (DMD), and two Limb Girdle 2A/R1 (LGMD2A/R1) patients on micromolded gelatin hydrogels. Engineered DMD and LGMD2A/R1 tissues demonstrated varying levels of defects in α-actinin expression and organization relative to control, depending on the mutation. In genetically relevant DMD tissues amenable to mRNA reframing by targeting exon 44 or 45 exclusion, exposure to exon skipping antisense oligonucleotides modestly increased myotube coverage and alignment and rescued dystrophin protein expression. These findings highlight the value of engineered culture substrates in guiding the organization of reprogrammed patient fibroblasts into aligned muscle tissues, thereby extending their value as tools for exploration and dissection of the cellular and molecular basis of genetic muscle defects, rescue, and repair.https://www.frontiersin.org/articles/10.3389/fcell.2022.830415/fullDMDLGMDexon skippinghydrogelscalpain 3dystrophin |
| spellingShingle | Florian Barthélémy Florian Barthélémy Jeffrey W. Santoso Laura Rabichow Laura Rabichow Rongcheng Jin Isaiah Little Isaiah Little Stanley F. Nelson Stanley F. Nelson Stanley F. Nelson Megan L. McCain Megan L. McCain M. Carrie Miceli M. Carrie Miceli Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels Frontiers in Cell and Developmental Biology DMD LGMD exon skipping hydrogels calpain 3 dystrophin |
| title | Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels |
| title_full | Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels |
| title_fullStr | Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels |
| title_full_unstemmed | Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels |
| title_short | Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels |
| title_sort | modeling patient specific muscular dystrophy phenotypes and therapeutic responses in reprogrammed myotubes engineered on micromolded gelatin hydrogels |
| topic | DMD LGMD exon skipping hydrogels calpain 3 dystrophin |
| url | https://www.frontiersin.org/articles/10.3389/fcell.2022.830415/full |
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