Fig. 6

Prolactin (PRL) reduces TNFα, IL-1β, and IFNγ (Cyt)-induced osteoclast formation in co-cultures of synovial fibroblast and osteoclast progenitor cells. a Time course of the co-culture: 12 hours after being seeded, synovial fibroblasts were treated or not with PRL for 16 hours. Cyt were then added or not to the culture and after 24 hours, osteoclast progenitors were delivered in medium containing 1, 25-dihydroxy-vitamin D3. Co-cultures were incubated for 10 days, replacing with new medium (containing +/- Cyt, +/- PRL, and 1, 25-dihydroxy-vitamin D3) every 2 days. Both synovial fibroblasts and osteoclast progenitors were from wild-type (Prlr+/+) mice. Osteclastogenesis, was assessed by the number of tartrate-resistant alkaline phosphatase (TRAP)-positive, multinucleated (≥3 nuclei) cells (TRAP(+) MNCs) (arrows). Scale bar 120 μm. b Number of TRAP(+) MNCs in co-cultures of synovial fibroblasts from Prlr+/+ mice and osteoclast progenitors from Prlr+/+ mice or from mice null for the PRL receptor (Prlr-/-). Co-cultures were carried out under the same conditions as in a Values are means ± SEM (n = 5). *P < 0.05, ***P < 0.001. c Real-time PCR amplification of the PRL receptor in joint cells obtained from Prlr+/+ and Prlr-/- mice. mRNA levels of Hprt were amplified as internal control. NC without RNA negative control