Ah Young Kim

Effect of Gelatin on Osteogenic Cell Sheet Formation Using Canine Adipose-Derived Mesenchymal Stem Cells.

Osteogenically differentiated cell sheet techniques using mesenchymal stem cells (MSCs) are available to stimulate bone regeneration. The advantage of the cell sheet technique is delivering live cells effectively into the focal region. We developed a novel osteogenic cell sheet technique by adding gelatin to osteogenic cell medium. Gelatin-induced osteogenic cell sheets (GCSs) were compared to conventional osteogenic cell sheets (OCSs). Undifferentiated MSCs (UCs) were used as a control. The morphology of these cell sheets was evaluated microscopically and histologically. The time-dependent cell proliferation rate was estimated by DNA quantification. The expression of osteogenic gene markers and the number of calcium depositions were assessed by quantitative real-time polymerase chain reaction and Alizarin red S (ARS) staining, respectively. GCSs were thicker and stronger than OCSs. GCSs showed a significantly higher cell proliferation rate compared to OCSs (p < 0.05). GCSs exhibited significantly higher upregulation of BMP-7 mRNA compared to OCSs (p < 0.05). Both GCSs and OCSs showed negative ARS reactivity on day 10, but only GCSs showed positive ARS reactivity on day 21. With this technique, we observed active cell proliferation with abundant ECM and upregulation of osteogenic bone markers, and our results suggest that GCSs could be promising for therapeutic applications in bone regeneration.

Comparison of viability and antioxidant capacity between canine adipose-derived mesenchymal stem cells and heme oxygenase-1-overexpressed cells after freeze-thawing.

Allogenic adipose-derived mesenchymal stem cells (Ad-MSCs) are an alternative source for cytotherapy owing to their antioxidant and anti-inflammatory effects. Frozen-thawed allogenic Ad-MSCs can be used instantly for this purpose. However, the viability and function of frozen-thawed Ad-MSCs have not been clearly evaluated. The purpose of this study was to compare the viability and function of Ad-MSCs and heme oxygenase-1 (HO-1)-overexpressed Ad-MSCs in vitro after freeze-thawing. The viability, proliferation, antioxidant capacity and mRNA gene expression of growth factors were evaluated. Frozen-thawed cells showed significantly lower viability than fresh cells (77% for Ad-MSCs and 71% for HO-1 Ad-MSCs, P<0.01). However, the proliferation rate of frozen-thawed Ad-MSCs increased and did not differ from that of fresh Ad-MSCs after 3 days of culture. In contrast, the proliferation rate of HO-1-overexpressed Ad-MSCs was lower than that of Ad-MSCs. The mRNA expression levels of TGF-β, HGF and VEGF did not differ between fresh and frozen-thawed Ad-MSCs, but COX-2 and IL-6 had significantly higher mRNA expression in frozen cells than fresh cells (P<0.05). Fresh Ad-MSCs exhibited higher HO-1 mRNA expression than frozen-thawed Ad-MSCs, and fresh HO-1-overexpressed Ad-MSCs exhibited higher than fresh Ad-MSCs (P<0.05). However, there was no significant difference between fresh and frozen HO-1-overexpressed Ad-MSCs. The antioxidant capacity of HO-1-overexpressed Ad-MSCs was significantly higher than that of Ad-MSCs. Cryopreservation of Ad-MSCs negatively affects viability and antioxidant capacity, and HO-1-overexpressed Ad-MSCs might be useful to maximize the effect of Ad-MSCs for cytotherapy.