Fig. 3.

Population doubling time (PDT) (a), TLR3 and TLR4 protein expression (b, c), and change in TLR3 and TLR4 mRNA expression (d) of OE-MSCs and AT-MSCs. The number of MSCs was counted following subculture from first passage and after calculation of average of cell count, the mean population doubling time was obtained according to the formula that is mentioned in “Materials and methods”. For evaluation of TLRs expression, MSCs were cultured to 70% confluence, harvested and then stained with the monoclonal antibodies against TLR3-PE and TLR4-PE. Isotype control was used as negative control. In addition, unspecific binding was removed by three time washing and by FC receptor blacking. The cells were then analyzed using flow cytometer. For TLR3 and TLR4 measurement at mRNA level, MSCs were grown to 70% confluence, total RNA was extracted and reverse transcribed. Real time was performed and the relative level of gene expression was calculated according to the 2^−ΔΔCT method. The experiment was performed in triplicate for each sample of OE-MSC and AT-MSC were in the same passage. Data shown represent mean ± SE of MSCs from 8 donners. *p < 0.05; **p < 0.01; ***p < 0.001. TLR toll like receptor