Ex vivo expansion and pluripotential differentiation of cryopreserved human bone marrow mesenchymal stem cells

Abstract

This study is aimed at investigating the potentials of ex vivo expansion and pluri-differentiation of cryopreservation of adult human bone marrow mesenchymal stem cells (hMSCs) into chondrocytes, adipocytes and neurocytes. Cryopreserved hMSCs were resuscitated and cultured for 15 passages, and then induced into chondrocytes, adipocytes and neurocytes with corresponding induction medium. The induced cells were observed for morphological properties and detected for expressions of type II collagen, triglyceride or neuron-specific enolase and nestin. The result showed that the resuscitated cells could differentiate into chondrocytes after exposure to transforming growth factor β 1 (TGF-β 1 ), insulin-like growth factor I (IGF-I) and vitamin C (V C ), and uniformly changed morphologically from a spindle-like fibroblastic appearance to a polygonal shape in three weeks. The induced cells were heterochromatic to safranin O and expressed cartilage matrix-procollagenal (II) mRNA. The resuscitated cells cultured in induction medium consisting of dexamethasone, 3-isobutyl-1-methylxanthine, indomethacin and IGF-I showed adipogenesis, and lipid vacuoles accumulation was detectable after 21 d. The resuscitated hMSCs were also induced into neurocytes and expressed nestin and neuron specific endolase (NSE) that were special surface markers associated with neural cells at different stage. This study suggested that the resuscitated hMSCs should be still a population of pluripotential cells and that it could be used for establishing an abundant hMSC reservoir for further experiment and treatment of various clinical diseases.

Cryopreservation characteristics of adipose-derived stem cells: maintenance of differentiation potential and viability

“…Overall, quantification of CFU frequencies of specific lineage phenotypes before and after freezing indicates that adipose-derived adult stem cells maintain their differentiation potential after cryopreservation. These findings are comparable to those reported for cryopreserved bone marrow-derived mesenchymal stem cells, which displayed comparable differentiation and proliferative potential preand post-thawing (Kotobuki et al, 2005;Xiang et al, 2007). Likewise, cryopreservation has been found not to alter the properties of synovial membrane-derived mesenchymal stem cells (De Bari et al, 2001).…”

Comparative studies of different cryopreservation methods for mesenchymal stem cells derived from human fetal liver

“…These findings suggest that the cryopreserved HFLCs still retain their multilineage differentiation potential. Other authors have reported similar observations for bone marrow-derived MSCs (Xiang et al, 2007). Furthermore, some investigators have shown that thawed human bone marrow MSCs were capable of tissue-engineered bone formation (Liu et al, 2008).…”

Use of Autologous Human mesenchymal Stromal Cell/Fibrin Clot Constructs in Upper Limb Non-Unions: Long-Term Assessment

“…MSC administration has been performed with two main procedures: ( i ) the “one-step” procedure, also known as “minimal manipulation”, using the bone marrow, either whole or concentrated, and ( ii ) the “two-step” procedure, also known as “extensive manipulation”, using ex vivo expanded and reimplanted bone marrow MSCs [41]. While the former method is simple and immediate, with an actual MSC concentration ranging in 1,000–10,000 cells/ml of bone marrow, the latter method is laborious and time-consuming, but it allows the MSC number to be increased by 100 to 10,000 folds [42].…”