gp130 in late osteoblasts and osteocytes is required for PTH-induced osteoblast differentiation

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   Figure 1

   Osteocytic gp130 is required for PTH to increase osteoblast numbers and bone formation in trabecular bone. Male mice were treated with hPTH (1–34) at 30 μg/kg per day for 5 weeks. (A) Numbers of osteoblasts/bone perimeter (N.Ob/B.Pm), (B) osteoblast surface/bone surface (Ob.S/BS), (C) osteoid surface/bone surface (OS/BS), (D) osteoid thickness (OTh), (E) double calcein-labeled surface (dLS/BS), and (F) mineral apposition rate (MAR) from trabecular bone in the proximal tibial secondary spongiosa in Dmp1Cre.gp130^w/w (gp130 w/w) and Dmp1Cre.gp130^f/f (gp130 f/f) mice. (G) The serum levels of procollagen type 1 amino-terminal propeptide (P1NP) and (H) endogenous murine PTH measured at the end of the treatment protocol are also shown. Scale bars are mean+s.e.m., n=8–10 per group. *P≤0.05, **P≤0.01 and ***P≤0.001, PTH-treated compared with genotype-matched vehicle-treated mice; ⁺⁺P<0.01 compared to vehicle-treated gp130 w/w mice.
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   Figure 2

   No effect of intermittent PTH on bone resorption. Male mice were treated with hPTH (1–34) at 30 μg/kg per day for 5 weeks. (A) Numbers of osteoclasts per unit bone perimeter (NOc/BPm), (B) numbers of osteoclasts per unit osteoclast perimeter (NOc/OcPm), (C) osteoclast surface per unit bone surface (OcS/BS) measured in the proximal tibial secondary spongiosa, and (D) serum levels of cross-linked C-terminal telopeptide of type 1 collagen (CTX1) in PTH and vehicle-treated Dmp1Cre.gp130^w/w (gp130 w/w) and Dmp1Cre.gp130^f/f (gp130 f/f) mice. Scale bars are mean+s.e.m., n=8–10 per group.
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   Figure 3

   PTH effects on cortical bone are impaired in Dmp1Cre.gp130^f/f mice. Male mice were treated with hPTH (1–34) at 30μg/kg per day for 5 weeks. (A) Tibial periosteal MAR (Ps.MAR), and (B) mineralizing surface per unit bone surface (MS/BS) in the tibial diaphysis, and (C) periosteal perimeter (Ps.Pm) of the femoral diaphysis in PTH and vehicle-treated Dmp1Cre.gp130^w/w (gp130 w/w) and Dmp1Cre.gp130^f/f (gp130 f/f) mice. Scale bars are mean+s.e.m., n=8–10 per group. **P≤0.01, NS, P>0.05 (not statistically significant) in PTH-treated compared with genotype-matched vehicle-treated mice. ⁺, P<0.05, vehicle-treated Dmp1Cre.gp130^f/f compared with vehicle-treated Dmp1Cre.gp130^w/w.
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   Figure 4

   PTH effects on WNT-inhibitor, but not osteoclastogenic, mRNA levels are impaired in DMP1Cre.gp130^f/f mice. RNA was isolated from femurs flushed of bone marrow and expression of PTH target genes was examined by relative quantitative PCR. Tnfs11 mRNA (A), Il6 mRNA (B), Dkk1 mRNA (C) and Sost mRNA (D) in Dmp1Cre.gp130^w/w and Dmp1Cre.gp130^f/f mice treated for 5 weeks with PTH, collected 1 h after the final injection. All values are shown relative to housekeeping (Hkg) – the geometric mean of hypoxanthine phosphoribosyltransferase 1 (Hprt1) and hydroxymethylbilane synthase (Hmbs). Scale bars are mean+s.e.m., n=5–8 bones per group, with mRNA prepared and analyzed separately. **P≤0.01 and ***P≤0.001, PTH-treated compared with genotype-matched vehicle-treated mice; ⁺P<0.05, vehicle-treated Dmp1Cre.gp130^f/f compared with vehicle-treated Dmp1Cre.gp130^w/w.
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   Figure 5

   PTH1R expression is reduced in DMP1Cre.gp130^f/f mice and gp130 deficient cultured osteoblasts. (A) Pth1r mRNA quantified by qPCR in femurs flushed of bone marrow obtained from untreated 12-week-old Dmp1Cre.gp130^w/w and Dmp1Cre.gp130^f/f mice, normalized to Hmbs; n=8 samples per group. (B) gp130 (Il6st) and (C) Pth1r, Runx2, Osx and Alpl mRNA levels in primary calvarial osteoblasts obtained from gp130^f/f or C57/BL6 WT neonates infected with lentiviral Cre-recombinase; levels are shown normalized to beta-2-microglobulin (B2m) (n=3 biological replicates). *P≤ 0.05; **P≤0.01, vs gp130 w/w or C57/BL6.

• © 2014 Society for Endocrinology