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

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Abstract
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.