master of molecular and developmental stem cell biology · 2012-11-30 · master of molecular and...
TRANSCRIPT
Master of Molecular and
Developmental Stem Cell Biology
Professor Dr. rer. nat. Dr. h. c. Beate Brand-Saberi
Department of Anatomy and Molecular Embryology
Institute of Anatomy, Faculty of Medicine
How do complex tissues
form from a few cells?
Mesodermal
stem cells Myoblasts
Myotubes,
Satellite cells Muscle fibers
Pax3
Pax7
Myf5
CXCR4
Myf5
MyoD
Mef2
Myf5
MyoD
Mef2
Myf5
MyoD
Mef2
Pax3
Pax7
Myf5
CXCR4
Mrf4
Myogenin
Pax7 (in satellite cells)
Muscle-specific proteins
Myf5
MyoD
Mef2
Myosin heavy chain
Desmin
Development of skeletal muscles
Paraxial mesoderm
somite
Somite sclerotome
and dermomyotome
Ventrolateral
dermomyotome limb
muscles
Development of skeletal muscles
Yusuf and Brand-Saberi, 2012
Wnt 1, 3a,4
Shh Noggin Fgf8
Wnt 6
BMP2,4
Intermediate Mesoderm Paraxial Mesoderm Neural Tube Notochord
BMP4
H.J. Jacob, Bochum/C. Hornik
Local signals determine the developmental fate
of somitic derivatives
Notch/
Delta
Vizualization of gene expression at mRNA-level by in situ hybridization
(Dr. Verena Chankiewitz)
MyoD is a bHLH transcription factor conserved during evolution
Nautilus in Drosophila
MyoD in vertebrates
NCBI: Probes for Functional Genomics
Brown C. In situ hybridization with riboprobes: an overview for veterinary pathologists.
Vet Pathol. 1998 May;35(3):159-67. PMID: 9598579
Satellite cell and its activation
Mitotically quiescent small mononuclear progenitor cells
Reside in a niche between the basal lamina and the sarcolemma of
their associated mature muscle fiber
Rudnicki, M A; et al. 2009
Modulpraktika im Schwerpunkt Molekulare
Biochemie der Stammzellen
• In situ Hybridisierung zur Darstellung der Expression von Entwicklungskontrollgenen im Vertebratenembryo
• Muskelstammzellen
und Myogenese
Abteilung für Anatomie und Molekulare Embryologie
Local signals determine the developmental fate
of somitic derivatives
Poland Syndrome is described as an
absence or underdevelopment of the
chest muscle (M. pectoralis) on one
side of the body and webbing of the
fingers (cutaneous syndactyly) of the
ipsilateral hand).
CXCR4/SDF-1 axis is involved in the migration of muscle precursor cells
into the limb bud
SDF-1:
Stromal-cell derived factor 1
Alpha-chemokine
CXCR4:
G protein-coupled receptor
SDF-1 is its sole ligand
Modulpraktikum in Wintersemester 2012/13 RUB
Isolation and in vitro differentiation
of murine embryonic stem cells
Dr. B. Wang & Dr. M. Napirei
Department of Anatomy and Molecular Embryology, RUB
Zygote Blastocyte
Cell culture
inner cell mass
ES cells
trophectoderm
blastocoel
Establishment of a murine embryonic stem (ES) cell line
A B
C D
Isolation and cultivation of blastocytes on mitotically arrested
murine embryonic fibroblasts (feeder cells)
Hatching of blastocyst from the zona pellucida and subsequent fast
upgrowth of the inner cell mass
Inner cell mass ready for trypsin treatment and transfer into the
24 well plate
Establishment of ES cell lines by picking out and culturing the colonies
separately
Establishment of a murine embryonic stem (ES) cell line
2 days old EB formed in the hanging-drop culture
4 days old EB in the static suspension culture
Differentiation of ES-cells by the embryoid body (EB) method
Culture of ES cells in hanging-drops
Formation of EBs in the hanging-drops
Culture of EBs in suspension medium
A B
C D
Isolation and cultivation of blastocytes on mitotically arrested murine embryonic
fibroblasts (feeder cells)
Hatching of blastocyst from the zona pellucida and subsequent fast upgrowth of the
inner cell mass (ES cells)
Inner cell mass ready for cell singling by trypsin treatment and
transfer on fresh feeder cells
Establishment of ES cell lines by picking and culturing separate ES cell
colonies
Establishment of a murine embryonic stem (ES) cell line
EB cells spontaneously differentiate into multipotent stem cells of the three germinal layers and subsequently to
terminally differentiated body cells
Abteilung für Anatomie und
Molekulare Embryologie
Klassische
Embryologie Entwicklungsbiologie
am Tiermodell
Hühnerembryo
Zebrafisch
Maus F
oto
: E
sth
er
Stö
ckli,
Zü
rich
Transmissions-
elektronenmikroskopie
Rasterelektronen-
mikroskopie
Molekularbiologie
Konfokalmikroskopie
Methoden