Abstract
OBJECTIVES:Articular cartilage is vulnerable to injuries and undergoes an irreversible degenerative process. The use of amniotic fluid mesenchymal stromal stem cells for the reconstruction of articular cartilage is a promising therapeutic alternative. The aim of this study was to investigate the chondrogenic potential of amniotic fluid mesenchymal stromal stem cells from human amniotic fluid from second trimester pregnant women in a micromass system (high-density cell culture) with TGF-β3 for 21 days.METHODS:Micromass was performed using amniotic fluid mesenchymal stromal stem cells previously cultured in a monolayer. Chondrocytes from adult human normal cartilage were used as controls. After 21 days, chondrogenic potential was determined by measuring the expression of genes, such as SOX-9, type II collagen and aggrecan, in newly differentiated cells by real-time PCR (qRT-PCR). The production of type II collagen protein was observed by western blotting. Immunohistochemistry analysis was also performed to detect collagen type II and aggrecan. This study was approved by the local ethics committee.RESULTS:SOX-9, aggrecan and type II collagen were expressed in newly differentiated chondrocytes. The expression of SOX-9 was significantly higher in newly differentiated chondrocytes than in adult cartilage. Collagen type II protein was also detected.CONCLUSION:We demonstrate that stem cells from human amniotic fluid are a suitable source for chondrogenesis when cultured in a micromass system. amniotic fluid mesenchymal stromal stem cells are an extremely viable source for clinical applications, and our results suggest the possibility of using human amniotic fluid as a source of mesenchymal stem cells.
Highlights
Chondrocytes represent the only cell type present in articular cartilage and are responsible for its homeostasis [1]
Amniocentesis was suggested by amniocentesis obstetrics upon suspicion of chromosomal abnormalities according to the Fetal Medicine-specific protocol, Hospital of Clinics, State University of Campinas (UNICAMP)
Amniotic fluid stem cell characterization Immunophenotypic characterization by flow cytometry revealed that mesenchymal stem cells (MSCs) from human amniotic fluid (HAF) were positive for mesenchymal markers CD90 (63.3%), CD73 (99.5%), and CD105 (22.7%)
Summary
Chondrocytes represent the only cell type present in articular cartilage and are responsible for its homeostasis [1]. The cartilage extracellular matrix (ECM) is composed of a network, including collagens, proteoglycans and other smaller components. Collagen represents approximately 70-80% of the dry tissue weight of cartilage and ensures its strength and structural organization. Aggrecan is the second most important component of the ECM, and it provides the mechanical properties that allow cartilage to be compressed [2]. Cartilage is known for its limited ability to repair or regenerate itself, which is due avascularity and a small.
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