Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Identification of critical molecular pathways involved in exosome-mediated improvement of cardiac function in a mouse model of muscular dystrophy


Duchenne muscular dystrophy (DMD) is a progressive disease characterized by skeletal muscle atrophy, respiratory failure, and cardiomyopathy. Our previous studies have shown that transplantation with allogeneic myogenic progenitor-derived exosomes (MPC-Exo) can improve cardiac function in X-linked muscular dystrophy (Mdx) mice. In the present study we explored the molecular mechanisms underlying this beneficial effect. We quantified gene expression in the hearts of two strains of Mdx mice (D2.B10-DmdMdx/J and Utrntm1Ked-DmdMdx/J). Two days after MPC-Exo or control treatment, we performed unbiased next-generation RNA-sequencing to identify differentially expressed genes (DEGs) in treated Mdx hearts. Venn diagrams show a set of 780 genes that were ≥2-fold upregulated, and a set of 878 genes that were ≥2-fold downregulated, in both Mdx strains following MPC-Exo treatment as compared with control. Gene ontology (GO) and protein-protein interaction (PPI) network analysis showed that these DEGs were involved in a variety of physiological processes and pathways with a complex connection. qRT-PCR was performed to verify the upregulated ATP2B4 and Bcl-2 expression, and downregulated IL-6, MAPK8 and Wnt5a expression in MPC-Exo-treated Mdx hearts. Western blot analysis verified the increased level of Bcl-2 and decreased level of IL-6 protein in MPC-Exo-treated Mdx hearts compared with control treatment, suggesting that anti-apoptotic and anti-inflammatory effects might be responsible for heart function improvement by MPC-Exo. Based on these findings, we believed that these DEGs might be therapeutic targets that can be explored to develop new strategies for treating DMD.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Fig. 1: Transcriptome analysis and DEG identification.
Fig. 2: Gene ontology (GO) analysis.
Fig. 3: The constructed PPI network of the upregulated and downregulated differentially expressed genes.
Fig. 4: qRT-PCR verification of the expression levels of 6 genes in Utrntm1Ked DmdMdx/J mice treated with PBS or MPC-Exo.
Fig. 5: Western blot verification of the protein levels of Bcl-2, IL-6, Wnt5a, and MAPK8 in Utrntm1Ked DmdMdx/J mice treated with PBS or MPC-Exo.


  1. 1.

    Chung J, Smith AL, Hughes SC, Niizawa G, Abdel-Hamid HZ, Naylor EW, et al. Twenty-year follow-up of newborn screening for patients with muscular dystrophy. Muscle Nerve. 2016;53:570–8.

    PubMed  Google Scholar 

  2. 2.

    Hoffman EP, Brown RH Jr, Kunkel LM. Dystrophin: the protein product of the Duchenne muscular dystrophy locus. Cell. 1987;51:919–28.

    CAS  Google Scholar 

  3. 3.

    Mauduit O, Delcroix V, Lesluyes T, Perot G, Lagarde P, Lartigue L, et al. Recurrent DMD Deletions Highlight Specific Role of Dp71 Isoform in Soft-Tissue Sarcomas. Cancers (Basel). 2019;11:922.

    CAS  Google Scholar 

  4. 4.

    Fukunaga H, Sonoda Y, Atsuchi H, Osame M. Respiratory failure and its care in Duchenne muscular dystrophy. Rinsho Shinkeigaku. 1991;31:154–8.

    CAS  PubMed  Google Scholar 

  5. 5.

    Kuno A, Horio Y. SIRT1: a novel target for the treatment of muscular dystrophies. Oxid Med Cell Longev. 2016;2016:6714686.

    PubMed  PubMed Central  Google Scholar 

  6. 6.

    Eagle M, Baudouin SV, Chandler C, Giddings DR, Bullock R, Bushby K. Survival in Duchenne muscular dystrophy: improvements in life expectancy since 1967 and the impact of home nocturnal ventilation. Neuromuscul Disord. 2002;12:926–9.

    PubMed  Google Scholar 

  7. 7.

    McNally EM, Kaltman JR, Benson DW, Canter CE, Cripe LH, Duan D, et al. Contemporary cardiac issues in Duchenne muscular dystrophy. Working Group of the National Heart, Lung, and Blood Institute in collaboration with Parent Project Muscular Dystrophy. Circulation. 2015;131:1590–8.

    PubMed  PubMed Central  Google Scholar 

  8. 8.

    Kamdar F, Garry DJ. Dystrophin-deficient cardiomyopathy. J Am Coll Cardiol. 2016;67:2533–46.

    CAS  PubMed  Google Scholar 

  9. 9.

    Su X, Jin Y, Shen Y, Ju C, Cai J, Liu Y, et al. Exosome-derived dystrophin from allograft myogenic progenitors improves cardiac function in duchenne muscular dystrophic mice. J Cardiovasc Transl Res. 2018;11:412–9.

    PubMed  PubMed Central  Google Scholar 

  10. 10.

    Su X, Shen Y, Jin Y, Jiang M, Weintraub N, Tang Y. Purification and transplantation of myogenic progenitor cell derived exosomes to improve cardiac function in duchenne muscular dystrophic mice. J Vis Exp. 2019.

  11. 11.

    Hrdlickova R, Toloue M, Tian B. RNA-Seq methods for transcriptome analysis. Wiley interdisciplinary reviews. RNA. 2017.

  12. 12.

    Ruan XF, Li YJ, Ju CW, Shen Y, Lei W, Chen C, et al. Exosomes from Suxiao Jiuxin pill-treated cardiac mesenchymal stem cells decrease H3K27 demethylase UTX expression in mouse cardiomyocytes in vitro. Acta Pharmacol Sin. 2018;39:579–86.

    CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Ruan XF, Ju CW, Shen Y, Liu YT, Kim IM, Yu H, et al. Suxiao Jiuxin pill promotes exosome secretion from mouse cardiac mesenchymal stem cells in vitro. Acta Pharmacol Sin. 2018;39:569–78.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. 14.

    Huang da W, Sherman BT, Lempicki RA. Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nat Protoc. 2009;4:44–57.

    PubMed  Google Scholar 

  15. 15.

    Zhou G, Soufan O, Ewald J, Hancock REW, Basu N, Xia J. NetworkAnalyst 3.0: a visual analytics platform for comprehensive gene expression profiling and meta-analysis. Nucleic Acids Res. 2019;47:W234–w41.

    CAS  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Schmittgen TD, Livak KJ. Analyzing real-time PCR data by the comparative C(T) method. Nat Protoc. 2008;3:1101–8.

    CAS  PubMed  Google Scholar 

  17. 17.

    Su X, Jin Y, Shen Y, Kim IM, Weintraub NL, Tang Y. RNAase III-type enzyme dicer regulates mitochondrial fatty acid oxidative metabolism in cardiac mesenchymal stem cells. Int J Mol Sci. 2019;20:5554.

    CAS  PubMed Central  Google Scholar 

  18. 18.

    Chen Z, Su X, Shen Y, Jin Y, Luo T, Kim IM, et al. MiR322 mediates cardioprotection against ischemia/reperfusion injury via FBXW7/notch pathway. J Mol Cell Cardiol. 2019;133:67–74.

    CAS  PubMed  PubMed Central  Google Scholar 

  19. 19.

    Li D, Long C, Yue Y, Duan D. Sub-physiological sarcoglycan expression contributes to compensatory muscle protection in mdx mice. Hum Mol Genet. 2009;18:1209–20.

    CAS  PubMed  PubMed Central  Google Scholar 

  20. 20.

    Bostick B, Yue Y, Long C, Duan D. Prevention of dystrophin-deficient cardiomyopathy in twenty-one-month-old carrier mice by mosaic dystrophin expression or complementary dystrophin/utrophin expression. Circ Res. 2008;102:121–30.

    CAS  PubMed  Google Scholar 

  21. 21.

    Rafael-Fortney JA, Chimanji NS, Schill KE, Martin CD, Murray JD, Ganguly R, et al. Early treatment with lisinopril and spironolactone preserves cardiac and skeletal muscle in Duchenne muscular dystrophy mice. Circulation. 2011;124:582–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Gardina PJ, Clark TA, Shimada B, Staples MK, Yang Q, Veitch J, et al. Alternative splicing and differential gene expression in colon cancer detected by a whole genome exon array. BMC Genomics. 2006;7:325.

    PubMed  PubMed Central  Google Scholar 

  23. 23.

    Cartwright EJ, Oceandy D, Austin C, Neyses L. Ca2+ signalling in cardiovascular disease: the role of the plasma membrane calcium pumps. Sci China Life Sci. 2011;54:691–8.

    CAS  PubMed  Google Scholar 

  24. 24.

    Sadi AM, Afroze T, Siraj MA, Momen A, White-Dzuro C, Zarrin-Khat D, et al. Cardiac-specific inducible overexpression of human plasma membrane Ca2+ ATPase 4b is cardioprotective and improves survival in mice following ischemic injury. Clin Sci (Lond). 2018;132:641–54.

    CAS  Google Scholar 

  25. 25.

    Lin B, Li Y, Han L, Kaplan AD, Ao Y, Kalra S, et al. Modeling and study of the mechanism of dilated cardiomyopathy using induced pluripotent stem cells derived from individuals with Duchenne muscular dystrophy. Dis Models Mechanisms. 2015;8:457–66.

    CAS  Google Scholar 

  26. 26.

    Brocheriou V, Hagege AA, Oubenaissa A, Lambert M, Mallet VO, Duriez M, et al. Cardiac functional improvement by a human Bcl-2 transgene in a mouse model of ischemia/reperfusion injury. J Gene Med. 2000;2:326–33.

    CAS  PubMed  Google Scholar 

  27. 27.

    Imahashi K, Schneider MD, Steenbergen C, Murphy E. Transgenic expression of Bcl-2 modulates energy metabolism, prevents cytosolic acidification during ischemia, and reduces ischemia/reperfusion injury. Circ Res. 2004;95:734–41.

    CAS  PubMed  Google Scholar 

  28. 28.

    Kirshenbaum LA, de Moissac D. The bcl-2 gene product prevents programmed cell death of ventricular myocytes. Circulation. 1997;96:1580–5.

    CAS  PubMed  Google Scholar 

  29. 29.

    Gustafsson AB, Gottlieb RA. Bcl-2 family members and apoptosis, taken to heart. Am J Physiol Cell Physiol. 2007;292:C45–51.

    CAS  PubMed  Google Scholar 

  30. 30.

    Abdel-Salam E, Abdel-Meguid I, Korraa SS. Markers of degeneration and regeneration in Duchenne muscular dystrophy. Acta Myol. 2009;28:94–100.

    CAS  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Pelosi L, Berardinelli MG, De Pasquale L, Nicoletti C, D’Amico A, Carvello F, et al. Functional and morphological improvement of dystrophic muscle by interleukin 6 receptor blockade. EBioMedicine. 2015;2:285–93.

    PubMed  PubMed Central  Google Scholar 

  32. 32.

    Smart N, Mojet MH, Latchman DS, Marber MS, Duchen MR, Heads RJ. IL-6 induces PI3-kinase and nitric oxide-dependent protection and preserves mitochondrial function in cardiomyocytes. Cardiovasc Res. 2006;69:164–77.

    CAS  PubMed  Google Scholar 

  33. 33.

    Gwechenberger M, Mendoza LH, Youker KA, Frangogiannis NG, Smith CW, Michael LH, et al. Cardiac myocytes produce interleukin-6 in culture and in viable border zone of reperfused infarctions. Circulation. 1999;99:546–51.

    CAS  PubMed  Google Scholar 

  34. 34.

    Hirota H, Izumi M, Hamaguchi T, Sugiyama S, Murakami E, Kunisada K, et al. Circulating interleukin-6 family cytokines and their receptors in patients with congestive heart failure. Heart Vessels. 2004;19:237–41.

    PubMed  Google Scholar 

  35. 35.

    Fontes JA, Rose NR, Cihakova D. The varying faces of IL-6: From cardiac protection to cardiac failure. Cytokine. 2015;74:62–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Coppe JP, Desprez PY, Krtolica A, Campisi J. The senescence-associated secretory phenotype: the dark side of tumor suppression. Annu Rev Pathol. 2010;5:99–118.

    CAS  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Ortiz-Montero P, Londono-Vallejo A, Vernot JP. Senescence-associated IL-6 and IL-8 cytokines induce a self- and cross-reinforced senescence/inflammatory milieu strengthening tumorigenic capabilities in the MCF-7 breast cancer cell line. Cell Commun Signal. 2017;15:17.

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  38. 38.

    Katsuumi G, Shimizu I, Yoshida Y, Minamino T. Vascular senescence in cardiovascular and metabolic diseases. Front Cardiovasc Med. 2018;5:18.

    PubMed  PubMed Central  Google Scholar 

  39. 39.

    Rose BA, Force T, Wang Y. Mitogen-activated protein kinase signaling in the heart: angels versus demons in a heart-breaking tale. Physiol Rev. 2010;90:1507–46.

    CAS  PubMed  Google Scholar 

  40. 40.

    Muslin AJ. MAPK signalling in cardiovascular health and disease: molecular mechanisms and therapeutic targets. Clin Sci (Lond). 2008;115:203–18.

    CAS  Google Scholar 

  41. 41.

    Petrich BG, Eloff BC, Lerner DL, Kovacs A, Saffitz JE, Rosenbaum DS, et al. Targeted activation of c-Jun N-terminal kinase in vivo induces restrictive cardiomyopathy and conduction defects. J Biol Chem. 2004;279:15330–8.

    CAS  PubMed  Google Scholar 

  42. 42.

    Siwik DA, Kuster GM, Brahmbhatt JV, Zaidi Z, Malik J, Ooi H, et al. EMMPRIN mediates beta-adrenergic receptor-stimulated matrix metalloproteinase activity in cardiac myocytes. J Mol Cell Cardiol. 2008;44:210–7.

    CAS  PubMed  Google Scholar 

  43. 43.

    Fan GC, Yuan Q, Song G, Wang Y, Chen G, Qian J, et al. Small heat-shock protein Hsp20 attenuates beta-agonist-mediated cardiac remodeling through apoptosis signal-regulating kinase 1. Circ Res. 2006;99:1233–42.

    CAS  PubMed  Google Scholar 

  44. 44.

    Anilkumar N, Sirker A, Shah AM. Redox sensitive signaling pathways in cardiac remodeling, hypertrophy and failure. Front Biosci (Landmark Ed). 2009;14:3168–87.

    CAS  Google Scholar 

  45. 45.

    Abraityte A, Vinge LE, Askevold ET, Lekva T, Michelsen AE, Ranheim T, et al. Wnt5a is elevated in heart failure and affects cardiac fibroblast function. J Mol Med (Berl, Ger). 2017;95:767–77.

    CAS  Google Scholar 

  46. 46.

    Abraityte A, Lunde IG, Askevold ET, Michelsen AE, Christensen G, Aukrust P, et al. Wnt5a is associated with right ventricular dysfunction and adverse outcome in dilated cardiomyopathy. Sci Rep. 2017;7:3490.

    PubMed  PubMed Central  Google Scholar 

  47. 47.

    Newman DR, Sills WS, Hanrahan K, Ziegler A, Tidd KM, Cook E, et al. Expression of WNT5A in idiopathic pulmonary fibrosis and its control by TGF-beta and WNT7B in human lung fibroblasts. J Histochem Cytochem. 2016;64:99–111.

    CAS  PubMed  Google Scholar 

  48. 48.

    Jin Y, Shen Y, Su X, Weintraub N, Tang Y. CRISPR/Cas9 technology in restoring dystrophin expression in iPSC-derived muscle progenitors. J Vis Exp. 2019.

Download references


This study was funded by grants HL124097 and HL126949 (NLW), HL134354 (YLT and NLW), and HL086555 (YLT) from the National Institutes of Health.

Author information




YLT designed the research; XS, YS, YJ performed the research; YLT and XS analyzed the data; and XS, NLW and YLT wrote the paper.

Corresponding author

Correspondence to Yao-liang Tang.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Su, X., Shen, Y., Jin, Y. et al. Identification of critical molecular pathways involved in exosome-mediated improvement of cardiac function in a mouse model of muscular dystrophy. Acta Pharmacol Sin 42, 529–535 (2021).

Download citation


  • Duchenne muscular dystrophy
  • cardiomyopathy
  • exosome
  • RNA-Seq
  • gene ontology
  • protein-protein interaction network
  • Bcl-2
  • IL-6

Further reading


Quick links