LACK OF EVIDENCE FOR AN INCREASE IN INTERLEUKIN-6 EXPRESSION IN ADULTMURINE BONE, BONE-MARROW, AND MARROW STROMAL CELL-CULTURES AFTER OVARIECTOMY

Interleukin-6 (IL-6) has been implicated as a mediator of postmenopaus al bone loss, In vitro studies of bone and bone marrow cells have sugg ested that estrogen regulates bone turnover by controlling the product ion of IL-6, a potent stimulator of osteoclastogenesis and bone resorp tion, To investigate this hypothesis in an in vivo model, we examined the effect of ovariectomy or estrogen replacement on IL-6 mRNA and pro tein expression in adult mouse bone and bone marrow in vivo and in mar row stromal cell cultures, Eight-week-old CD-1 mice were sham-operated (SHAM), ovariectomized (OVX), or ovariectomized and subcutaneously im planted with 21-day slow-release pellets containing 10 mu g of 17 beta -estradiol (O+E), Placebo pellets were implanted in the SHAM and OVX m ice. Uterine weights at 1, 2, or 3 weeks after surgery were significan tly decreased (68-76%) in OVX animals compared with SHAM or O+E. In mi ce sacrificed at 1 or 3 weeks after surgery, we found by nonquantitati ve reverse transcribed polymerase chain reaction (RT-PCR), that SHAM, OVX, and O+E calvariae (CALV) constitutively expressed IL-6 mRNA. In c ontrast, IL-6 mRNA was either barely detectable or absent in the tibia (TIB) and bone marrow (BM). In the mice sacrificed 3 weeks after surg ery, we determined by quantitative RT-PCR that IL-6 mRNA in the CALV f rom the OVX and O+E groups were decreased by 81 and 92%, respectively, compared with SHAM. IL-6 protein levels in the pushed bone marrow (BM Sups) were detectable and were not significantly different among the g roups. In bone marrow cells that were cultured for 1 week, basal level s of IL-6 protein were low and did not differ significantly among the SHAM, OVX,or O+E groups sacrificed 1, 2, or 3 weeks after surgery, Aft er the addition of hrIL-1 alpha, IL-6 protein levels increased 100- to 1300-fold over control. IL-6 levels in cells from animals sacrificed 2 weeks after surgery were significantly lower in the hrIL-1 alpha-sti mulated OVX and O+E groups than in hrIL-1 alpha-stimulated SHAM cell c ultures. In conclusion, in this model we could find no increase in IL- 6 production with in vivo estrogen withdrawal in calvaria, long bones, bone marrow, or marrow stromal cell cultures, If increases in IL-6 ex pression are involved in the effects of estrogen withdrawal on bone, t he magnitude of these changes are relatively small and below the limit s of detection of the assays that we employed.