Osteogenic differentiation of intact human amniotic membrane.
Tissue engineering strategies usually require cell isolation and combination with a suitable biomaterial. Human amniotic membrane (AM) represents a natural two-layered sheet comprising cells with proven stem cell characteristics. In our approach, we evaluated the differentiation potential of AM in toto with its sessile stem cells as alternative to conventional approaches requiring cell isolation and combination with biomaterials. For this, AM-biopsies were differentiated in vitro using two osteogenic media compared with control medium (CM) for 28 days. Mineralization and osteocalcin expression was demonstrated by (immuno)histochemistry. Alkaline phosphatase (AP) activity, calcium contents and mRNA expression of RUNX2, AP, osteopontin, osteocalcin, BMP-2 (bone morphogenetic protein), and BMP-4 were quantified and AM viability was evaluated. Under osteogenic conditions, AM-biopsies mineralized successfully and by day 28 the majority of cells expressed osteocalcin. This was confirmed by a significant rise in calcium contents (up to 27.4 ± 6.8 mg/dl d28), increased AP activity, and induction of RUNX2, AP, BMP-2 and BMP-4 mRNA expression. Relatively high levels of viability were retained, especially in osteogenic media (up to 78.3 ± 19.0% d14; 62.9 ± 22.3% d28) compared to CM (42.2 ± 15.2% d14; 35.1 ± 8.6% d28). By this strategy, stem cells within human AM can successfully be driven along the osteogenic pathways while residing within their natural environment.
Lindenmair
Andrea
A
Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Research Center, Donaueschingenstrasse 13, A-1200 Vienna, Austria.
Wolbank
Susanne
S
Stadler
Guido
G
Meinl
Alexandra
A
Peterbauer-Scherb
Anja
A
Eibl
Johann
J
Polin
Helene
H
Gabriel
Christian
C
van Griensven
Martijn
M
Redl
Heinz
H
D000469:Alkaline Phosphatase / Q000378:metabolism; D000650:Amnion / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D015415:Biomarkers / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003470:Culture Media / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D042541:Intracellular Space / Q000187:drug effects / Q000201:enzymology; D010012:Osteogenesis / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction
D015415:Biomarkers; D003470:Culture Media; D000469:Alkaline Phosphatase; D002118:Calcium