Periodic Heat Shock Accelerated the Chondrogenic Differentiation of Human Mesenchymal Stem Cells in Pellet Culture

Abstract

Osteoarthritis (OA) is one of diseases that seriously affect elderly people's quality of life. Human mesenchymal stem cells (hMSCs) offer a potential promise for the joint repair in OA patients. However, chondrogenic differentiation from hMSCs in vitro takes a long time (∼6 weeks) and differentiated cells are still not as functionally mature as primary isolated chondrocytes, though chemical stimulations and mechanical loading have been intensively studied to enhance the hMSC differentiation. On the other hand, thermal stimulations of hMSC chondrogenesis have not been well explored. In this study, the direct effects of mild heat shock (HS) on the differentiation of hMSCs into chondrocytes in 3D pellet culture were investigated. Periodic HS at 41°C for 1 hr significantly increased sulfated glycosaminoglycan in 3D pellet culture at Day 10 of chondrogenesis. Immunohistochemical and Western Blot analyses revealed an increased expression of collagen type II and aggrecan in heat-shocked pellets than non heat-shocked pellets on Day 17 of chondrogenesis. In addition, HS also upregulated the expression of collagen type I and X as well as heat shock protein 70 on Day 17 and 24 of differentiation. These results demonstrate that HS accelerated the chondrogenic differentiation of hMSCs and induced an early maturation of chondrocytes differentiated from hMSCs. The results of this study will guide the design of future protocols using thermal treatments to facilitate cartilage regeneration with human mesenchymal stem cells.

Species-specific developmental timing is maintained by pluripotent stem cells ex utero

“…In another model, differentiation of pancreatic beta cells was also sped up compared to all other previous in vitro protocols (Pagliuca et al, 2014). Because the timing of many embryonic events during human development is often not known accurately due to the lack of embryonic material, it is often unclear from these studies whether developmental timing was altered per se, or if the apparent acceleration was due to improving conditions such that the rate of differentiation better approximated the actual in vivo rate (Chen et al, 2014, Ozeki et al, 2012, Tadeu et al, 2015). Whether exposing human pluripotent stem cell to a mouse embryonic environment can more dramatically accelerate differentiation is currently unclear.…”

Inhibition of microRNA-222 expression accelerates bone healing with enhancement of osteogenesis, chondrogenesis, and angiogenesis in a rat refractory fracture model

“…To form pellet cultures, 2.5 Â 10 5 hMSCs were centrifuged (1200 rpm, 5 min) in a 15 mL Falcon Tube (Corning, NY, USA). Cells were cultured under chondrogenic conditions for 14 days in differentiation medium (Differentiation Basal Medium-Chondrogenic, LONZA) with 1% antibiotic-antimycotic solution, 37 nM BMP2 (Wako Pure Chemical Industries, Ltd. Osaka Japan) and 0.79 nM TGF-b3 (Wako Pure Chemical Industries, Ltd.) according to a previously described protocol [11]. The pellet medium was changed every 3 days, and cell pellets were cultured for 14 days with chondrogenic medium prior to analysis.…”

Culture temperature affects redifferentiation and cartilaginous extracellular matrix formation in dedifferentiated human chondrocytes

“…However, most in vitro studies on chondrocytes or AC have been performed using a culture temperature of 37°C, which may not accurately reflect the in vivo temperature. In addition, the effect of a high‐temperature environment, such as 41°C, remains unclear, although an intermittent heat stimulus (41°C) has been reported to potentially have a positive effect on ECM formation …”