Periosteum as a source of mesenchymal stem cells: the effects of TGF-β3 on chondrogenesis

INTRODUCTION: Numerous experimental efforts have been undertaken to induce the healing of lesions within articular cartilage by re-establishing competent repair tissue. Adult mesenchymal stem cells have attracted attention as a source of cells for cartilage tissue engineering. The purpose of this study was to investigate chondrogenesis employing periosteal mesenchymal cells. METHODS: Periosteum was harvested from patients who underwent orthopedic surgeries. Mesenchymal stem cells were characterized through flow cytometry using specific antibodies. The stem cells were divided into four groups. Two groups were stimulated with transforming growth factor β3 (TGF-β3), of which one group was cultivated in a monolayer culture and the other was cultured in a micromass culture. The remaining two groups were cultivated in monolayer or micromass cultures in the absence of TGF-β3. Cell differentiation was verified through quantitative reverse transcription-polymerase chain reaction (RT-PCR) and using western blot analysis. RESULT: In the groups cultured without TGF-β3, only the cells maintained in the micromass culture expressed type II collagen. Both the monolayer and the micromass groups that were stimulated with TGF-β3 expressed type II collagen, which was observed in both quantitative RT-PCR and western blot analysis. The expression of type II collagen was significantly greater in the micromass system than in the monolayer system. CONCLUSION: The results of this study demonstrate that the interactions between the cells in the micromass culture system can regulate the proliferation and differentiation of periosteal mesenchymal cells during chondrogenesis and that this effect is enhanced by TGF-β3.

Repair of critical-size defects with autogenous periosteum-derived cells combined with bovine anorganic apatite/collagen: an experimental study in rat calvaria

The aim of this study was to evaluate the bone repair using autogenous periosteum-derived cells (PDC) and bovine anorganic apatite and collagen (HA-COL). PDC from Wistar rats (n=10) were seeded on HA-COL discs and subjected to osteoinduction during 6 days. Critical-size defects in rat calvarias were treated with blood clot (G1), autogenous bone (G2), HA-COL (G3) and HA-COL combined with PDC (G4) (n=40), and then analyzed 1 and 3 months after surgeries. Radiographic analysis exhibited no significant temporal change. G1 and G2 had discrete new marginal bone, but the radiopacity of graft materials in G2, G3 and G4 impaired the detection of osteogenesis. At 3 months, histopathological analysis showed the presence of ossification islets in G1, which was more evident in G2, homogeneous new bone around HA-COL in G3 and heterogeneous new bone around HA-COL in G4 in addition to moderate presence of foreign body cells in G3 and G4. Histomorphometric analysis showed no change in the volume density of xenograft (p>0.05) and bone volume density in G2 was twice greater than in G1 and G4 after 3 months (p<0.05), but similar to G3. The PDC did not increase bone formation in vivo, although the biomaterial alone showed biocompatibility and osteoconduction capacity.

O objetivo deste estudo foi avaliar o reparo ósseo usando células derivadas de periósteo (PDC) e apatita inorgânica e colágeno bovinos (HA-COL). PDC de ratos Wistar (n=10) foram semeadas sobre discos de HA-COL e osteoinduzidas por 6 dias. Defeitos de tamanho crítico em calvárias de ratos foram tratados com coágulo sanguíneo (G1), osso autógeno (G2), HA-COL (G3) ou HA-COL associado a PDC (G4) (n=40) e analisados em 1 e 3 meses após as cirurgias. Análise radiográfica não exibiu mudança temporal significante, G1 e G2 tiveram aumento discreto de novo osso marginal, entretanto a radiopacidade dos materiais de enxerto em G2, G3 e G4 prejudicou a detecção de osteogênese. Análise histopatológica mostrou em 3 meses ilhotas de ossificação em G1 que foi maior em G2, novo osso homogêneo ao redor de HA-COL em G3 e novo osso heterogêneo ao redor de HA-COL em G4 além da presença moderada de células gigantes de corpo estranho em G3 e G4. Análise histomorfométrica mostrou a densidade de volume inalterada do xenoenxerto (p>0,05) e a densidade de volume de novo osso em G2 duas vezes maior que G1 e G4 após 3 meses (p<0,05), mas similar a G3. PDC não aumentaram a formação óssea in vivo apesar do biomaterial sozinho ter apresentado biocompatibilidade e capacidade osteocondutora.

Strategy for generating tissue-engineered human bone construct

The strategy used to generate tissue-engineered bone construct, in view of future clinical application is presented here. Osteoprogenitor cells from periosteum of consenting scoliosis patients were isolated. Growth factors viz TGF-B2, bFGF and IGF-1 were used in concert to increase cell proliferation during in vitro cell expansion. Porous tricalcium phosphate (TCP)-hydroxyapatite (HA) scaffold was used as the scaffold to form 3D bone construct. We found that the addition of growth factors, greatly increased cell growth by 2 to 7 fold. TCP/HA proved to be the ideal scaffold for cell attachment and proliferation. Hence, this model will be further carried out on animal trial.

Autologous human fibrin as the biomaterial for tissue engineering

Patient own fibrin may act as the safest, cheapest and immediate available biodegradable scaffold material in clinical 1 tissue engineering. This study investigated the feasibility of using patient own fibrin isolated from whole blood to construct a new human cartilage, skin and bone. Constructed in vitro tissues were implanted on the dorsal part of the nude mice for in vivo maturation. After 8 weeks of implantation, the engineered tissues were removed for histological analysis. Our results demonstrated autologous fibrin has great potential as clinical scaffold material to construct various human tissues.