Equine marrow-derived mesenchymal stem cells: isolation, differentiation and culture optimization

Most studies regarding the marrow-derived equine mesenchymal stem cells [MSCs] have mainly focused on the cell transplantation without considering the capacity of differentiation and in vitro requirements of the cells. These concerns were investigated in the present study. Equine MSCs were isolated from the sternal marrow aspirates and expanded through two successive subcultures. Passage-2 equine MSC cultures were then treated with appropriate supplements in order to examine the cell osteogenic, chondrogenic and adipogenic differentiation potential. Furthermore, the culture of the cells was investigated in terms of the optimal concentration of fetal bovine serum [FBS] and the initial cell-seeding density. Additionally, a growth curve was plotted for the cells to study their growth characteristics. According to our findings, equine MSCs were easily generated specialized bone, cartilage and adipose cell lineages as confirmed by specific staining and RT-PCR analysis. Moreover, the cells exhibited rapid expansion when being cultivated in the medium with 15% FBS at 100 cells/cm[2]. Growth curves indicated that these cells rapidly entered the log phase after a brief lag [adaptation] period. In summary, marrow-derived equine MSCs possess tripotent differentiation capacity and rapid growth rate in the appropriate culture conditions

[Effects of lithium chloride on in vitro proliferation rate of rat marrow-derived mesenchymal stem cells]

To evaluate the effects of lithium chloride on MSCs in vitro expansion rate. In this experimental study, bone marrow from 8 rats was plated at 5x105-cells/cm2 in the presence of 1,2,5,7 and 10 mM Lithium chloride and expanded through 3 passages. Twelve days after initiatial culture, the cells of different groups were stained with crystal violet in order to compare the number and diameter of the colonies. Also the cells from different groups were compared in terms of the population doubling [PD] during the 1-3 passages. Different groups of growth curves were plotted for the third passage cells. At the end of cultivation period, the cells were examined wheatear they could differentiate into bone and adipose cells. The Number and diameter of the colonies in primary cultures treated with 5mM lithium chloride were significantly higher than those of control and other groups [P<0.001]. The cell population in the culture with 5 mM lithium chloride was doubled in average 12.02 +/- 0.04 times during 1-3 passages that was significantly higher than other groups. Compared to other groups, the cells from 5 mM group were reached platue in a short time [4.9 days] [P<0.001]. Alizarin red staining for bone and oil red for adipose cells indicated that the cells in different studied groups preserved their differentiation potential. Finally, it seems that the presence of 5 mM litium chloride in the cultures of rat bone marrow cells enhances the MSC in vitro expansion rate while maintaining their bone and adipose differentiation potential