Induction of chondro-, osteo- and adipogenesis in embryonic stem cells by bone morphogenetic protein-2: effect of cofactors on differentiating lineages.
BACKGROUND
Recently, tissue engineering has merged with stem cell technology with interest to develop new sources of transplantable material for injury or disease treatment. Eminently interesting, are bone and joint injuries/disorders because of the low self-regenerating capacity of the matrix secreting cells, particularly chondrocytes. ES cells have the unlimited capacity to self-renew and maintain their pluripotency in culture. Upon induction of various signals they will then differentiate into distinctive cell types such as neurons, cardiomyocytes and osteoblasts.


RESULTS
We present here that BMP-2 can drive ES cells to the cartilage, osteoblast or adipogenic fate depending on supplementary co-factors. TGFbeta1, insulin and ascorbic acid were identified as signals that together with BMP-2 induce a chondrocytic phenotype that is characterized by increased expression of cartilage marker genes in a timely co-ordinated fashion. Expression of collagen type IIB and aggrecan, indicative of a fully mature state, continuously ascend until reaching a peak at day 32 of culture to approximately 80-fold over control values. Sox9 and scleraxis, cartilage specific transcription factors, are highly expressed at very early stages and show decreased expression over the time course of EB differentiation. Some smaller proteoglycans, such as decorin and biglycan, are expressed at earlier stages. Overall, proteoglycan biosynthesis is up-regulated 7-fold in response to the supplements added. BMP-2 induced chondrocytes undergo hypertrophy and begin to alter their expression profile towards osteoblasts. Supplying mineralization factors such as beta-glycerophosphate and vitamin D3 with the culture medium can facilitate this process. Moreover, gene expression studies show that adipocytes can also differentiate from BMP-2 treated ES cells.


CONCLUSIONS
Ultimately, we have found that ES cells can be successfully triggered to differentiate into chondrocyte-like cells, which can further alter their fate to become hypertrophic, and adipocytes. Compared with previous reports using a brief BMP-2 supplementation early in differentiation, prolonged exposure increased chondrogenic output, while supplementation with insulin and ascorbic acid prevented dedifferentiation. These results provide a foundation for the use of ES cells as a potential therapy in joint injury and disease.
zur Nieden
Nicole I
NI
Molecular & Genetic Toxicology, Bayer HealthCare AG, Wuppertal, Germany. nicole.zur.nieden@gmx.net
Kempka
Grazyna
G
Rancourt
Derrick E
DE
Ahr
Hans-Jürgen
HJ
D017667:Adipocytes / Q000166:cytology; D050156:Adipogenesis; D053674:Aggrecans; D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology; D051792:Basic Helix-Loop-Helix Proteins / Q000096:biosynthesis / Q000235:genetics; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology*; D002356:Cartilage / Q000166:cytology; D002454:Cell Differentiation; D002460:Cell Line; D019902:Chondrocytes / Q000166:cytology; D020219:Chondrogenesis; D003094:Collagen / Q000096:biosynthesis / Q000235:genetics; D004622:Embryo, Mammalian / Q000166:cytology; D016326:Extracellular Matrix Proteins / Q000096:biosynthesis / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D006609:High Mobility Group Proteins / Q000096:biosynthesis / Q000235:genetics; D007328:Insulin / Q000494:pharmacology; D037181:Lectins, C-Type / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D010006:Osteoblasts / Q000166:cytology; D010012:Osteogenesis; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000502:physiology; D011506:Proteins / Q000494:pharmacology; D011509:Proteoglycans / Q000096:biosynthesis / Q000235:genetics; D055758:SOX9 Transcription Factor; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D053674:Aggrecans; D001205:Ascorbic Acid; D051792:Basic Helix-Loop-Helix Proteins; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D003094:Collagen; D016326:Extracellular Matrix Proteins; D006609:High Mobility Group Proteins; D007328:Insulin; D037181:Lectins, C-Type; D011506:Proteins; D011509:Proteoglycans; D055758:SOX9 Transcription Factor; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; C508320:Acan protein, mouse; C525719:Bmp2 protein, mouse; C503815:Scx protein, mouse; C527933:Sox9 protein, mouse