In addition, COLII-coated microbeads resulted in hypertrophic maturation of the differentiated chondrocytes, similar to conventional pellet culture, while CS-coated microbeads were able to retain the pre-hypertrophy state of the differentiated cells.132 Those findings demonstrated that CS coatings are beneficial to induce MSCs to differentiate into chondrocytes and prevent hypertrophy. Decellularized matrix ECM, an indispensable niche for stem cells microenvironment for stem cell rejuvenation during expansion.48, 49, 137, 138 Cartilage matrix A porous scaffold derived from adult porcine articular cartilage has the ability to induce chondrogenic differentiation of human ADSCs without exogenous growth factors, with significant synthesis and accumulation of ECM macromolecules, and with the development of mechanical properties approaching those of native cartilage.139 In a further study, acellular cartilage matrix (ACM) powders and human SDSCs were mixed into collagen gel for culture. and potential mechanisms underlying these strategies are also categorized. This paper aims to provide guidelines for the prevention of hypertrophy in the regeneration of cartilage tissue. This knowledge may also facilitate the retardation of osteophytes in the treatment of osteoarthritis. expansion without running the risk of losing their phenotype; however, MSCs tend to simultaneously acquire hypertrophic properties during chondrogenic induction, indicating the possibility of further differentiation toward endochondral bone formation.7, 8 It is becoming crucial to systematically assess current strategies for minimizing hypertrophy of chondrogenically differentiated cells to provide a high-quality cartilage tissue for clinical defect repair. A previous review covered molecular and biophysical mechanisms regulating hypertrophic differentiation in chondrocytes and MSCs9; this review will focus on strategies for preventing chondrogenic hypertrophy, including some new findings, such as the impact of different MSC sources and culture substrates. Potential mechanisms underlying Pico145 the above strategies will also be delineated. Pico145 Definition and characterization of chondrogenic hypertrophy Chondrogenic hypertrophy is marked by a more than 10-fold increase in cell volume and ECM Rabbit Polyclonal to Bax (phospho-Thr167) structural remodeling.10 Cell volume expansion affects cell function.11 The explosive increase in the volume of hypertrophic chondrocytes involves changes in intracellular and extracellular osmolarity, ECM degradation around the cell, and an increase in the amount of organelles per cell.12 Osmotic swelling has been shown stereologically to be responsible for most of the cell volume increase. Swelling can be the result of either an increase in cytoplasmic concentration or a decrease in extracellular osmolarity followed by aquaporin-mediated movement of water to re-establish iso-osmotic conditions.13 Of all the ECM molecules, AGC is the prime contributor to the osmotic pressure generated in cartilage, both due to its abundance and its high negative fixed charge. It is not completely understood if expression of terminal markers results in increased cell volume or vice versa. Chondrocyte hypertrophic differentiation is the gradual development process from chondrogenic differentiation to Pico145 cartilage mineralization, which is characterized by a series of markers; each of these markers has its own function in the process of cartilage mineralization.14 For example, the transcription factors, runt-related transcription factor 2 (RUNX2) and myocyte enhancer factor-2C (MEF2C), drive the expression of terminal differentiation markers, including matrix metalloproteinase 13 (MMP13),9 collagen type X (COLX),15 Indian hedgehog (IHH),16 alkaline phosphatase (ALP), and vascular endothelial growth factor (VEGF),8, 17 which all functionally contribute to endochondral ossification. Secreted MMP13 degrades COLII and AGC, key ECM components of functional cartilage18; COLX serves as a framework for subsequent calcification through matrix vesicles (MV)19; ALP hydrolyses pyrophosphate (PPi) to inorganic phosphate (Pi) which, in the presence of calcium, forms hydroxyapatite20; and IHH induces the proliferation of non-hypertrophic chondrocytes.21 Calcification of cartilage ECM originates at MV.22 ECM mineralization to endochondral bone formation consists of three steps (Fig.?1): (1) Hydroxyapatite crystals are formed inside the MV; (2) Hydroxyapatite crystals penetrate MV into the ECM; and (3) Endochondral ossification. The final stages of endochondral ossification, including degradation of the calcified matrix, VEGF-mediated vascular invasion of the calcified zone, and deposition of osteoid on the calcified trabeculae by osteoblasts, Pico145 are all under the control of MMPs.23 MMP is indispensable for the development of MV and it can calcify the growth plate; finally, calcification is substituted by endochondral bone. MMP13 binding to the MV membrane and cooperating with MMP9 could promote the release of VEGF in apoptotic chondrocytes, further accelerating the formation of vascularity in the growth plate.24 Open in a separate window Figure?1 ECM mineralization process: (1) Hydroxyapatite crystals are formed inside the MV (gray shading) when the concentration of calcium ion (influx through annexinII/V/VI calcium ion channels) and Pi [produced by the hydrolysis of Pcho and PEA via PHOSPHO1236, 237 and transferred into the MV by type-III Na+/Pi cotransporter238, 239 exceeds the solubility values.20, 238 (2) Hydroxyapatite.