Manifestation of Novel Social Challenges of the European Unionin the Teaching Material ofMedical Biotechnology Master’s Programmesat the University of Pécs and at the University of DebrecenIdentification number: TÁMOP-4.1.2-08/1/A-2009-0011

REGENERATION AND TRANSDIFFERENTIATION OF SKELETAL MUSCLE

Dr. Péter Balogh and Dr. Péter EngelmannTransdifferentiation and regenerative medicine – Lecture 7

Manifestation of Novel Social Challenges of the European Unionin the Teaching Material ofMedical Biotechnology Master’s Programmesat the University of Pécs and at the University of DebrecenIdentification number: TÁMOP-4.1.2-08/1/A-2009-0011

TÁMOP-4.1.2-08/1/A-2009-0011

Conditions requiring skeletal muscle regenerationInjury leading to extensive muscle damageInherited diseases – Duchenne’s muscular dystrophy:• X-linked mutation of dystrophin gene• 1:3500 males affected• Dystrophin (2.4 Mb in size) is the largest known

mammalian gene • Onset of the disease: DMD-afflicted patients are

diagnosed in childhood. The progressive muscle-wasting disease affects striated muscle including limb muscles, diaphragm, and heart leading to cardiorespiratory failure, and death usually occurs in the teenage years or early 20s.

TÁMOP-4.1.2-08/1/A-2009-0011Experimental models for studying muscle regeneration• Mdx mice: spontaneous mutation of the distrophin

gene (variable severities in different inbred mouse strains)

• Distrophin/utrophin double mutant mouse• Canine X-linked muscular dystrophy (cxmd) is the

best representation of DMD, but the phenotype is variable.

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Embyonic development of skeletal muscle

Myf5 Myf6Pax3

MyoD

MyogenesisMyogn Myf6, MyoD

NT

NC

MTSC

Limb VLL

DTDT

SC

Myf5Pax3/Pax7

MyoD

Bmp4

Wnt1/3Nog

Wnt11

Pax3

MyoD ShhWnt7a

NogMyf5

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Transcriptional control of myogenic differentiation

Transit Amplifying cells

MyotubeMyoblastDifferentiationActivation/Proliferation

Myogenic progenitor cells (MPC)Myogenic stem cell (MSC)Quiescent

Cd34Cdh15Foxk1MetPax3Pax7Sox8Sdc4Sox15Vcam1

Myf5Myf6MyoD

DesMyog

Myofibernuclei

Injury Fusion Differentiation Maturation

Proliferation andself-renewal of satellite cells

Regeneratingmyofiber nuclei

Satellite cell(quiescent)

Satellite cell(quiescent)

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Cellular sources for muscle regeneration• Satellite cells and their precursors• Endothelial cells associated with embryonic limb

muscles• Mesangioblasts• Bone marow-derived stem cells • Pluripotent cells found within muscle-derived side

population (SP) cells• Highly active Mdr-dependent expulsion of Hoechst

33342 dye

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Tissue sources for muscle regeneration

Vascular progenitors

Interstitial cells

Bone marrow cells

Myofibernuclei

Satellitecell

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Muscle stem cells – satellite cells• The satellite cells reside beneath the basal lamina of

muscle, closely juxtaposed to muscle fibers• Approximately up 2–7% of the nuclei associated with

a particular fiber• Heterogeneous composition: fusing/non-fusing

subsets• Ontogeny: somite/perivascular cells expressing

Pax3/Pax7• Surface markers

– Mouse: M-cadherin, CD34, VCAM, CD56, c-met (HGF-receptor)

– Human: CD56

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Structure and regeneration of skeletal muscle

Myofibril

Hematopoietic cells

Pericyte

Endothelial cell

Arteriole andcapillaries

Interstitial cell

Basal lamina

Satellite cell(SC)

Muscle fiber

Myonucleus

Quiescent SCPax7+

Activated SCPax7+

Myf5+MyoD+

Fusion anddifferentiation

Return toquiescence

MyoblastPax7-

Myf5+MyoD+

Expansion(symmetric

division)

Asymmetricdivision

Activation

MyocyteMyoD+

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Kinetics of muscle repair

Activation

Proliferation

Differentiation

Maturation

0 1 2 5 10 14Days post injury

TÁMOP-4.1.2-08/1/A-2009-0011Problems with myoblast regeneration in Duchenne’s muscular distrophy• Necessity for immunosuppression • Immunosuppressant drugs cause myoblast apoptosis• Short migratory distance following intramuscular

injection – 100 injections/cm2 (totalling up to 4,000 injections in a single patient!)

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Non-SCs contributing to muscle regeneration

Expansion

Commitment (if needed) Allogeneic transplantationAutologous transplantation(after genetic correction)

Mesenchymal differentation

Adipose-derived stem cellsMyoD-converted cells

HSCsSide population

Mesenchymal stem cellsMAPCs

SCs and subpopulationsMDSCs

CD133+ stem cells

HSCsSide population

CD133+ stem cells

MABs/pericytesMyoendothelial cells

EPCsMSCs

iPS cells

Reprogramming

Dermis or other tissues

Skeletal muscle

Bone marrow

Other sources

Blood

Vessels

Characterization

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Summary

• The prime candidates for skeletal muscle regeneration are the satellite cells, but cells from other sources (embryonic as well as non-embryonic) may also associate/promote the process.

• Muscle regeneration is accomplished through (a) promoting vascular repair, (b) cellular differentiation from muscle stem cells and (c) possible transdifferentiation.