Osteogenic Differentiation of hASCs and hBMMSCs

hASCs and hBMMSCs were seeded on different surfaces as described above, in 6-well plates. After 7 and 14 days of osteoinduction, the Trizol reagent (Invitrogen, Carlsbad, CA, USA) was used to isolate the total cellular RNAs of each group. After synthesizing the first strand cDNA using the reverse transcription system (Roche, Basel, Switzerland), quantification of all gene transcripts was performed by real-time polymerase chain reaction (qPCR) using a Power SYBR Green PCR Master Mix and an ABI PRISM 7500 sequence detection system (Applied Biosystems, Foster City, CA, USA). The β-actin expression was used as the internal control. The primer sequences were shown in Table 1. The cycle threshold values (Ct values) were used to calculate the fold differences among the samples, using the ∆∆Ct method^{65 (link),66 (link)}.Table 1

Primers for realtime PCR.

Gene	Forward primers	Reverse primers
RUNX2	ATGGGATGGGTGTCTCCACA	CCACGAAGGGGAACTTGTC
OSX	CCTCCTCAGCTCACCTTCTC	GTTGGGAGCCCAAATAGAAA
OCN	CACTCCTCGCCCTATTGGC	CCCTCCTGCTTGGACACAAAG
β-actin	CATGTACGTTGCTATCCAGGC	CTCCTTAATGTCACGCACGAT