Wada S, Martin T J, Findlay D M
St. Vincent's Institute of Medical Research, Fitzroy, Australia.
Endocrinology. 1995 Jun;136(6):2611-21. doi: 10.1210/endo.136.6.7750484.
Calcitonin (CT) down-regulates its receptor in several cancer cell lines, including T47D human breast cancer cells. Removal of CT results in the recovery of CT receptor (CTR) binding. However, homologous regulation of the CTR in osteoclasts is not well understood. To elucidate these phenomena in cells of the osteoclast lineage, mouse osteoblasts and bone marrow cells were cocultured on type 1 collagen gels. For the experiments, osteoclast-like cell (OCL)-enriched populations were subcultured from the collagen gels into multiwell dishes on days 7-8, and CT regulation of the CTR was determined and compared with that in T47D cells. When cells of either type were treated with CT for 1 h and then washed, binding capacity for [125I] salmon CT ([125I]sCT) was decreased dependent upon the preincubating concentration of CT. After removal of CT, the binding capacity in OCLs recovered toward the control level over 12 h. However, in contrast to that in T47D cells, recovery was transient, so that 24 h after removal of CT, the binding capacity in preincubated cells was strikingly reduced compared with that in control cells. This occurred even when the preincubating concentration of CT was too low to cause down-regulation of binding in 1 h. Scatchard analysis showed a decrease in receptor number in CT-treated compared with control OCLs 24 h after CT removal, with unchanged receptor affinity. By autoradiography, decreased CTR density on multinuclear OCLs was indicated. Preexposure of either OCLs or T47D cells to CT caused elevation of intracellular cAMP, which persisted for 6-12 h after removal of CT. In addition, there was desensitization to a rechallenge with CT, which, in T47D cells, recovered by 24-36 h. In contrast, OCLs showed incomplete recovery of desensitization. These data correlated with the results of semiquantitative reverse transcription-polymerase chain reaction studies; the CTR messenger RNA level was increased to about 150% of the control level in sCT-treated T47D cells 18-36 h after sCT removal; the level was markedly decreased to about 20% of the control value in sCT-treated OCLs 12-48 h after sCT removal and remained suppressed. This study suggests that CT-induced homologous down-regulation is a potential cause of the "escape" phenomenon, by producing a population of CT-resistant osteoclasts.
降钙素（CT）可下调包括T47D人乳腺癌细胞在内的多种癌细胞系中的其受体。去除CT后，降钙素受体（CTR）结合能力得以恢复。然而，破骨细胞中CTR的同源调节作用尚不清楚。为了阐明破骨细胞谱系细胞中的这些现象，将小鼠成骨细胞和骨髓细胞在I型胶原凝胶上共培养。在实验中，在第7 - 8天将富含破骨细胞样细胞（OCL）的细胞群体从胶原凝胶中传代培养至多孔培养皿中，并测定CT对CTR的调节作用，并与T47D细胞中的情况进行比较。当两种类型的细胞用CT处理1小时后冲洗，[125I]鲑鱼降钙素（[125I]sCT）的结合能力会根据CT的预孵育浓度而降低。去除CT后，OCL中的结合能力在12小时内恢复至对照水平。然而，与T47D细胞不同的是，这种恢复是短暂的，因此在去除CT 24小时后，预孵育细胞中的结合能力与对照细胞相比显著降低。即使CT的预孵育浓度过低以至于在1小时内不会导致结合能力下调时，这种情况仍会发生。Scatchard分析显示，在去除CT 24小时后，与对照OCL相比，CT处理的OCL中受体数量减少，而受体亲和力不变。通过放射自显影显示，多核OCL上的CTR密度降低。将OCL或T47D细胞预先暴露于CT会导致细胞内cAMP升高，在去除CT后这种升高持续6 - 12小时。此外，对再次用CT刺激会产生脱敏现象，在T47D细胞中，这种脱敏现象在24 - 36小时后恢复。相比之下，OCL的脱敏恢复不完全。这些数据与半定量逆转录 - 聚合酶链反应研究结果相关；在去除sCT 18 - 36小时后，sCT处理的T47D细胞中CTR信使RNA水平增加至对照水平的约150%；在去除sCT 12 - 48小时后，sCT处理的OCL中该水平显著降低至对照值的约20%并持续受到抑制。这项研究表明，CT诱导的同源下调是产生一群CT抗性破骨细胞从而导致“逃逸”现象的潜在原因。
