Figure 6.

Effect of FGF23 on PTH secretion (a and b) and expression of cell proliferation markers (c,d and e) in cultured parathyroid glands. A parathyroid gland with some attached thyroid tissue was micro-dissected from mice under a stereomicroscope, coated with collagen gel, and incubated in MEM with FGF23 (100 ng/mL) or vehicle (saline) for 4 days as described in the Methods section. (a) Time-course of the effect of FGF23 on PTH secretion from normal parathyroid glands. The amount of PTH secreted after 1- and 13-h incubation with (black bar) or without (white bar) FGF23 (100 ng/mL) is expressed as a cumulative value, while the value on day 4 represents the amount of PTH secreted in 24 h. PTH secreted in the culture medium versus baseline medium PTH secretion in the initial 2-h incubation period was regarded as representing PTH secretion under the specific condition. (b) Effect of FGF23 on PTH secretion on day 4 in the parathyroid glands from mice with or without cKO of Fgfr1–3 or αKlotho. Fgfr1–3^flox/flox and αKlotho^flox/flox mice were mated with PTH-Cre mice (cKO, black bar) or not mated (non-cKO, white bar). PTH secretion experiments were carried out as described above. (c) Percentage of Ki67+ cell number in cultured parathyroid glands. The Ki67+ and DAPI+ cell numbers were detected and counted as described in the Methods section. (d) Representative micrographs of Ki67 expression (red) with DAPI staining (blue) for nuclei in tissue sections of cultured parathyroid glands. (e) Representative micrographs of staining for phosphorylated ERK1/2 (pERK; green) in parathyroid tissue sections. Scale bars: 10 μm. Values are expressed as mean ± s.d. *P < 0.05, **P < 0.01, ***P < 0.001. NS, not significantly different (P > 0.05).