Characterization of primary calvaria cultures from TR4−/−vs. TR4+/+mice. A: Proliferation rate was compared between primary calvaria cultures of TR4−/− and TR4+/+ pups by 3 H-thymidine incorporation assay. The 20 K, 50 K, and 100 K numbers represent different cell number seeding density. B: Quantification of osteoblastic marker gene expression. Total RNA were harvested from the primary calvaria cells of TR4−/− and TR4+/+ mice, and converted to first strand cDNA, and subjected to q-PCR. The relative expression fold of the genes from TR4−/− mice, as compared to TR4+/+ mice, were calculated. C: The serum levels of osteocalcin from TR4−/− and TR4+/+ mice were determined by ELISA.