Mineralized Human Amniotic Membrane as a Biomimetic Scaffold for Hard Tissue Engineering Applications.

The human amniotic membrane (HAM) has been viewed as a potential regenerative material for a wide variety of injured tissues because of its collagen-rich content. High degradability of HAM limits its wide practical application in bone tissue engineering. In this study, the natural matrix of the decellularized amniotic membrane was developed by the double diffusion method. The results confirmed a reduction of the amniotic membrane's degradability because of the deposition of calcium and phosphate ions during the double diffusion process. Real-time PCR results showed a high expression of osteogenesis-related genes from adipose-derived mesenchymal stem cells (ADMSCs) cultured on the surface of the developed mineralized amniotic membrane (MAM). Further in vivo experiments were conducted using an MAM preseeded with ADMSCs and a critical-size rat calvarial defect model. Histopathological results confirmed that the MAM + cell sample has excellent potential in bone regeneration.

Platelet-rich plasma application in chondrogenesis.

Platelet-rich plasma (PRP), an autologous derivative of whole blood, has been recently used in surgical treatment. PRP contains growth factors including transforming growth factor-ß (TGF-ß), insulin-like growth factor (IGF), platelet-derived growth factor (PDGF), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) and also bioactive proteins that influence the healing of tendon, ligament, muscle, and bone. This article describes the current clinical applications of PRP in chondrogenesis. This study reviews and evaluates the studies that have been published in the field of chondrogenesis. All aspects of using PRP in chondrogenesis are reviewed.

Assessment of TGF-ß3 on production of aggrecan by human articular chondrocytes in pellet culture system.

BACKGROUND: The Autologous Chondrocytes Transplantation (ACT) method is being studied for repair of cartilage diseases. As the chondrocytes dedifferentiated during monolayer culture, three-dimensional cultures are suggested to redifferentiate them. The aim of this study was investigation of the effect of TGF-ß3 growth factor on chondrocytes in pellet culture system. MATERIALS AND METHODS: The chondrocytes were isolated from three human articular cartilages by enzymatic digestion. The cells of the second passage were transferred to pellet culture system. We determined the chondrogenic medium with TGF-ß3 as the experimental group and without it as the control group. After 2 weeks, the aggrecan production was investigated using histological and immunohistochemical (IHC) methods. RESULTS: The presence of glycosaminoglycans was proved through Toluiden blue staining. Comparison of IHC results using MATLAB software showed that aggrecan in the experimental group was significantly higher than in the control group (P ≤ 0.05). CONCLUSION: The presence of TGF-ß3 in the chondrogenic medium could lead to the production of more aggrecan in chondrocytes cultivated in pellet culture system.

The effect of platelet rich plasma on chondrogenic differentiation of human adipose derived stem cells in transwell culture.

OBJECTIVE(S): Platelet-rich plasma (PRP) has recently emerged as a promising strategy in regenerative medicine due to its multiple endogenous growth factors. Little is known about the role of PRP as a promoter in chondrogenesis of human adipose derived stem cells (hADSCs). The aim of this study was to determine whether PRP may be considered as a natural and easy achievable source of growth factors to promote the chondrogenic differentiation of hADSCs in Transwell culture. Materials and Methods : Biochemical, immunohistological and molecular assays were used to evaluate the effect of different concentrations (5%, 10%, and 15%) of PRP on chondrogenic differentiation of hADSCs in Transwell culture. Results : The cells in the presence of 10% PRP produced markedly higher amounts of GAG and DNA, in comparison to the control group. PRP also increased chondrogenic markers in these cells, such as sox-9, aggrecan and collagen type II. A high expression level of collagen type X as a hypertrophic marker was observed in cartilage produced by using either PRP or TGF-ß1. Conclusion : Our findings indicate that autologous PRP at an optimum concentration had beneficial effects on differentiation of hADSCs in Transwell culture. Further, in vivo studies are necessary to fully define the clinical implications of PRP.

A comparative study of aggrecan synthesis between natural articular chondrocytes and differentiated chondrocytes from adipose derived stem cells in 3D culture.

INTRODUCTION: The main obstacle for tissue engineering is to find the most appropriate cell which is able to produce extracellular matrix (ECM) similar or better than natural chondrocytes in vitro. This study compared aggrecan synthesis's potential between differentiated chondrocytes (DCs) from adipose-derived stem cells (ADSCs) and natural articular chondrocytes (NCs) in 3D culture in vitro. MATERIALS AND METHODS: Human ADSCs were isolated from sub-cutaneous adipose tissue and then the surface markers including CD 14, 45 CD105, CD90, CD44 were analyzed by flow cytometry. Also human articular chondrocytes were yielded of non-weight bearing area of Knee cartilage. Both types of the cells were encapsulated in alginate scaffolds and cultured in chondrogenic medium with and without TGFß3 for 3 weeks. Then the extent of aggercan (AGC) production was evaluated by ELISA on days 14 and 21. RESULTS: Our findings indicated that differentiated chondrocytes (DCs) with and without TGFß3 synthesized more AGC than natural chondrocytes (NCs) on day 14. But DCs without TGFß3 had higher production than other groups on day 21. Application of TGFß3 resulted in an increase of amount of AGC in DCs on day 14 but a decrease on day 21 than same group. CONCLUSION: Since, aggrecan is an important chondrogenic marker, it was concluded that ADSCs can be possible reliable alternative cell source for cartilage tissue engineering in future.

Effects of FGF-2 on human adipose tissue derived adult stem cells morphology and chondrogenesis enhancement in Transwell culture.

Injured cartilage is difficult to repair due to its poor vascularisation. Cell based therapies may serve as tools to more effectively regenerate defective cartilage. Both adult mesenchymal stem cells (MSCs) and human adipose derived stem cells (hADSCs) are regarded as potential stem cell sources able to generate functional cartilage for cell transplantation. Growth factors, in particular the TGF-b superfamily, influence many processes during cartilage formation, including cell proliferation, extracellular matrix synthesis, maintenance of the differentiated phenotype, and induction of MSCs towards chondrogenesis. In the current study, we investigated the effects of FGF-2 on hADSC morphology and chondrogenesis in Transwell culture. hADSCs were obtained from patients undergoing elective surgery, and then cultured in expansion medium alone or in the presence of FGF-2 (10 ng/ml). mRNA expression levels of SOX-9, aggrecan and collagen type II and type X were quantified by real-time polymerase chain reaction. The morphology, doubling time, trypsinization time and chondrogenesis of hADSCs were also studied. Expression levels of SOX-9, collagen type II, and aggrecan were all significantly increased in hADSCs expanded in presence of FGF-2. Furthermore FGF-2 induced a slender morphology, whereas doubling time and trypsinization time decreased. Our results suggest that FGF-2 induces hADSCs chondrogenesis in Transwell culture, which may be beneficial in cartilage tissue engineering.