Murine osteoblast interleukin 4 receptor expression: upregulation by 1,25 dihydroxyvitamin D3

The immune cytokine interleukin 4 has newly recognized effects on skeletal metabolism. While the interaction of many cells ultimately determines bone mass, we have examined the possibility that the osteoblast may be an IL-4 target in bone by characterizing IL-4 receptor (IL-4R) expression by MC3T3-E1 (MC3T3) murine osteoblastic cells. Based on 125I-IL-4 binding, MC3T3 cells express large numbers of IL-4 receptors (125I-IL-4 Bmax = 3,000-7,500 sites/cell, 125I-IL-4 K = 13-40 pM) with an affinity similar to the IL-4 receptor expressed by an IL-4-responsive T cell line. Monoclonal anti-IL-4R antibodies (M1) blocked specific MC3T3 125I-IL-4 binding and MC3T3 total cell RNA contained full-length IL-4R mRNA as detected by reverse transcription DNA amplification utilizing IL-4R primers and Northern blot analysis. Functionally, IL-4 treatment of MC3T3 cells resulted in increased cellular proliferation (10-20%) and inhibition of alkaline phosphatase levels (20-40%). While parathyroid hormone (PTH) exposure did not influence IL-4R levels, vitamin D3 treatment augmented MC3T3 125I-IL-4 binding, in a time-dependent manner, up to threefold after a 24 h exposure with a metabolite specificity indicating the involvement of the vitamin D receptor. Equilibrium binding studies showed that the impact of 1,25 (OH)2 D3 on MC3T3 125I-IL-4 binding was due to an increased IL-4R Bmax. Cycloheximide treatment inhibited 1,25 (OH)2 D3-induced IL-4R upregulation, suggesting that protein synthesis was required. Furthermore, the steroid increased steady-state IL-4R mRNA levels in both a time- and concentration-dependent manner. The IL-4R message half-life was not altered by 1,25 (OH)2 D3, suggesting that increased IL-4R mRNA expression resulted from increased IL-4R gene transcription. Taken together, these findings raise the possibility that IL-4's influence on mineral metabolism could be mediated by osteoblasts and that the effectiveness of this cytokine may be influenced by vitamin D3's impact on IL-4R expression.