e. The thermal cycling profile involved an initial activation for 10 minutes at 95˚C, followed by 15 seconds at 95˚C, 1 minute at 60˚C, and running for 40 cycles. The melting curve stage was set at 95˚C (15 seconds), 60˚C (1 minute), and 95˚C (15 seconds) ([@B22]). Gene expression values were calculated using the following formula:

ΔΔCT= \[minimum CT Targets-minimum CT RPL-13A\]Test samples-\[minimum CT Targets- minimum CT RPL-13A\]Stem cells

Real-time PCR was performed to compare the effects of 0.1 and 0.2 mT-fields on the expressions of the osteogenic markers. In another part of this study the effects of three daily-exposure durations of 2, 4 and 6 hours for electromagnetic field application were studied after 10 days of stimulation. Real-time PCR was used to compare gene expression levels among the above mentioned groups.

Surgical procedures
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Each animal was anesthetized and a small incision was made on the left side of the cranium. The periosteum and soft tissues were removed to access the cranial bone. The collagen-based scaffolds ([@B23]) with dimensions of 5×5×1 mm^3^were implanted after a predrilling with a dental drill. The following three groups were defined and studied in triplicate: i. Bone sockets without any scaffolds; ii. Defects filled with scaffolds seeded with untreated MSCs, and iii. Defects filled with scaffolds seeded with electromagnetically and chemically motivated MSCs. Vicryl 3-0 suture was used to close the incisions. We performed autologous transplantation and each animal used its own stem cells. The scaffolds were retrieved after 10 weeks of implantation and fixed with 4% paraformaldehyde for 12 hours. Thereafter, decalcification of bones was carried out in 10% EDTA for two weeks followed by embedding in paraffin. Tissue blocks were sectioned into 5 µm thick sections and stained with hematoxylin and eosin (H&E) to assess bone healing ([@B24]).

Statistical analysis
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All data that corresponded to the three separate experiments were 